Filamin a binding anti-inflammatory and analgesic

ABSTRACT

A compound or its pharmaceutically acceptable salt, optionally including both individual enantiomeric forms, a racemate, diastereomers and mixtures thereof, composition and method are disclosed that can provide analgesia and reduce inflammation. A contemplated compound has a structure that corresponds to Formula A, wherein the R group substituents, d, e, f, k, n, m, D, E, F, K, G, P, Q, W, and Z are defined within.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a continuation-in-part of application Ser. No.12/610,091 filed on Oct. 30, 2009 that claims priority from applicationSer. No. 12/435,304 that was filed on May 4, 2009, as well as Ser. No.12/263,257 that was filed on Oct. 31, 2008, whose disclosures areincorporated herein by reference.

TECHNICAL FIELD

This invention contemplates a composition and related method forproviding long-lasting analgesia and reducing inflammation. Moreparticularly, a compound, composition and method are described thatutilize a small molecule to bind filamin A, to reduce inflammation andto preserve Gi/o signaling by the mu opioid receptor, known to interactwith filamin A. Preferably, the compound reduces inflammation, preservesmu opioid receptor −Gi/o signaling and also functions as a mu opioidreceptor agonist. Most preferably, the compound binds filamin A withpicomolar or sub-picomolar affinity.

BACKGROUND OF THE INVENTION

Best known for cross-linking cytoplasmic actin into dynamic scaffolds tocontrol cell motility, filamins are large cytoplasmic proteinsincreasingly found to regulate cell signaling by interacting with over30 different receptors and signaling molecules (Feng et al., 2004 NatCell Biol 6:1034-1038; Stossel et al., 2001 Nature 2:138-145), includingthe mu opioid receptor (MOR) (Onoprishvili et al, 2003 Mol Pharmacol64:1092-1100). Filamins are dimerized through the last carboxy-terminalrepeat near the transmembrane regions, allowing an intracellularV-shaped structure that is critical for function. There are threemammalian isoforms: filamin A (FLNA), B and C.

FLNA controls cell motility by controlling the cycle of actinpolymerization and depolymerization, allowing cells to move and tomigrate. As actin depolymerization is linked to the inflammatoryresponse, binding to FLNA suppresses inflammation by slowing actinpolymerization and cell motility. Femtomolar naloxone and its inactiveisomer, both known to bind FLNA (Wang et al., 2008 PLoS One 3:e1554),have been shown to reduce the microglial inflammatory response; i.e.,pro-inflammatory factors and reactive oxygen species, oflipopolysaccaride-activated microglial cells (Liu et al, 2000 JPET293:607-617; Qin et al., 2005 FASEB J 19:550-557). The glialinflammatory response has been implicated in neuropathic pain(Hutchinson et al., 2008 Eur J Neurosci 28:20-29) as well as theinflammatory neurotoxicity of neurodegenerative disease (Liu et al.,2003 JPET 304:1-7).

A second function of binding to FLNA is a beneficial regulation ofopioid receptor signaling; i.e., a maintenance of coupling to Gi and Goproteins. MOR preferentially couples to pertussis toxin-sensitive Gproteins, Gi/o (inhibitory/other), and inhibits the adenylylcyclase/cAMP pathway (Laugwitz et al., 1993 Neuron 10:233-242; Connor etal., 1999 Clin Exp Pharmacol Physiol 26:493-499). Analgesia results fromthese MOR-linked inhibitory G protein (Gi/o) signaling cascades andrelated ion channel interactions that suppress cellular activities byhyperpolarization.

Adaptive responses of opioid receptors contribute to the development ofanalgesic tolerance and physical dependence, and possibly also tocomponents of opioid addiction. A critical adaptive response of the MORis a switch in G protein coupling from its native Gi/o proteins tostimulatory Gs proteins, resulting in opposite effects on the cell uponactivation as well as analgesic tolerance and physical dependence (Wanget al., 2005 Neuroscience 135:247-261). Prevention of this G proteincoupling switch by agents that bind filamin A (Wang et al, 2008 PLoS One3:e1554), a scaffolding protein known to interact with MOR, canalleviate unwanted adaptive responses to continued opioidadministration.

A chronic opioid-induced switch to Gs coupling by MOR can causeexcitatory signaling, by activation of adenylyl cyclase, in place of theusual inhibitory signaling or inhibition of adenylyl cyclase (Crain etal., 1992 Brain Res 575:13-24; Crain et al., 2000 Pain 84:121-131;Gintzler et al., 2001 Mol Neurobiol 21:21-33; Wang et al., 2005Neuroscience 135:247-261). This switch in G protein coupling from Gi/oto Gs (Wang et al., 2005 Neuroscience 135:247-261; Chakrabarti et al.,2005 Mol Brain Res 135:217-224) may be a result of the decreasedefficiency of coupling to the native G proteins, the usual index ofdesensitization (Sim et al., 1996 J Neurosci 16:2684-2692) and stillcommonly considered the reason for analgesic tolerance.

The chronic opioid-induced MOR-G protein coupling switch is accompaniedby stimulation of adenylyl cyclase II and IV by MOR-associated Gβγdimers (Chakrabarti et al., 1998 Mol Pharmacol 54:655-662; Wang et al.,2005 Neuroscience 135:247-261). The interaction of the Gβγ dimer withadenylyl cyclase had previously been postulated to be the sole signalingchange underlying the excitatory effects of opiates (Gintzler et al.,2001 Mol Neurobiol 21:21-33). It has further been shown that the Gβγthat interacts with adenylyl cyclases originates from the Gs proteincoupling to MOR and not from the Gi/o proteins native to MOR (Wang etal., 2006 J Neurobiol 66:1302-1310).

Thus, MORs are normally inhibitory G protein-coupled receptors thatcouple to Gi or Go proteins to inhibit adenylyl cyclase and decreaseproduction of the second messenger cAMP, as well as to suppress cellularactivities via ion channel-mediated hyperpolarization. Opioid analgesictolerance and dependence are also associated with that switch in Gprotein coupling by MOR from Gi/o to Gs (Wang et al., 2005 Neuroscience135:247-261). This switch results in activation of adenylyl cyclase thatprovides essentially opposite, stimulatory, effects on the cell.

Controlling this switch in G protein coupling by MOR is the scaffoldingprotein FLNA, and compounds that bind a particular segment of FLNA withhigh affinity, like naloxone (NLX) and naltrexone (NTX), can preventthis switch (Wang et al, 2008 PLoS One 3:e1554) and the associatedanalgesic tolerance and dependence (Wang et al., 2005 Neuroscience135:247-261). This switch in G protein coupling also occurs acutely,though transiently, and is potentially linked to the acute rewarding oraddictive effects of opioid drugs, through CREB activation as a resultof increased cAMP accumulation (Wang et al., 2009 PLoS ONE 4(1):e4282).

Ultra-low-dose NLX or NTX have been shown to enhance opioid analgesia,minimize opioid tolerance and dependence (Crain et al., 1995 Proc NatlAcad Sci USA 92:10540-10544; Powell et al. 2002. JPET 300:588-596), aswell as to attenuate the addictive properties of opioids (Leri et al.,2005 Pharmacol Biochem Behav 82:252-262; Olmstead et al., 2005Psychopharmacology 181:576-581). An ultra-low dose of opioid antagonistwas an amount initially based on in vitro studies of nociceptive dorsalroot ganglion neurons and on in vivo mouse studies. It has long beenhypothesized that ultra-low-dose opioid antagonists enhance analgesiaand alleviate tolerance/dependence by blocking the excitatory signalingopioid receptors that underlie opioid tolerance and hyperalgesia (Crainet al., 2000 Pain 84:121-131). Later research has shown that theattenuation of opioid analgesic tolerance, dependence and addictiveproperties by ultra-low-dose, defined herein, naloxone or naltrexone,occurs by preventing the MOR-Gs coupling that results from chronicopiate administration (Wang et al., 2005 Neuroscience 135:247-261), andthat the prevention of MOR-Gs coupling is a result of NLX or NTX bindingto filamin A at approximately 4 picomolar affinity (Wang et al, 2008PLoS One 3:e1554).

Found in all cells of the brain, CREB is a transcription factorimplicated in addiction as well as learning and memory and several otherexperience-dependent, adaptive (or maladaptive) behaviors (Carlezon etal., 2005 Trends Neurosci 28:436-445). In general, CREB is inhibited byacute opioid treatment, an effect that is completely attenuated bychronic opioid treatment, and activated during opioid withdrawal(Guitart et al., 1992 J Neurochem 58:1168-1171). However, a regionalmapping study showed that opioid withdrawal activates CREB in locuscoeruleus, nucleus accumbens and amygdala but inhibits CREB in lateralventral tegemental area and dorsal raphe nucleus (Shaw-Luthman et al.,2002 J Neurosci 22:3663-3672).

In the striatum, CREB activation has been viewed as a homeostaticadaptation, attenuating the acute rewarding effects of drugs (Nestler,2001 Am J Addict 10:201-217; Nestler, 2004 Neuropharmacology 47:24-32).This view is supported by nucleus accumbens overexpression of CREB or adominant-negative mutant respectively reducing or increasing therewarding effects of opioids in the conditioned place preference test(Barot et al., 2002 Proc Natl Acad Sci USA 99:11435-11440). In conflictwith this view, however, is the finding that reducing nucleus accumbensCREB via antisense attenuated cocaine reinforcement as assessed inself-administration (Choi et al., 2006 Neuroscience 137:373-383).Clearly, CREB activation is implicated in addiction, but whether itdirectly contributes to the acute rewarding effects of drugs orinitiates a homeostatic regulation thereof appears less clear.

The several-fold increase in pS¹³³CREB reported by Wang et al., 2009PLoS ONE 4(1):e4282 following acute, high-dose morphine may indicateacute dependence rather than acute rewarding effects. However, thetransient nature of the MOR-Gs coupling correlating with this CREBactivation suggests otherwise. In fact, the correlation of pS¹³³CREBwith the Gs coupling by MOR following this acute high-dose morphineexposure, as well as the similar treatment effects on both, suggest thatthis alternative signaling mode of MOR can contribute to the acuterewarding or addictive effects of opioids. This counterintuitive notioncan explain the apparent paradox that ultra-low-dose NTX, whileenhancing the analgesic effects of opioids, decreases the acuterewarding or addictive properties of morphine or oxycodone as measuredin conditioned place preference or self-administration and reinstatementparadigms.

In considering analgesic tolerance, opioid dependence, and opioidaddiction together as adaptive regulations to continued opioid exposure,a treatment that prevents MOR's signaling adaptation of switching its Gprotein partner can logically attenuate these seemingly divergentbehavioral consequences of chronic opioid exposure.

Even though ultra-low-dose NTX blocks the conditioned place preferenceto oxycodone or morphine (Olmstead et al., 2005 Psychopharmacology181:576-581), its co-self-administration only reduces the rewardingpotency of these opioids but does not abolish self-administrationoutright (Leri et al., 2005 Pharmacol Biochem Behav 82:252-262). It ispossible that a direct stimulatory effect on VTA neurons, as opposed tothe proposed disinhibition via inhibition of GABA interneurons (Spanagelet al., 1993 Proc Natl Acad Sci USA 89:2046-2050), can play some role inopioid reward. A MOR-Gs coupling mediation of reward, increasing withincreasing drug exposure, is in keeping with current theories that theescalation of drug use signifying drug dependence can not indicate a“tolerance” to rewarding effects but instead a sensitization torewarding effects (Zernig et al., 2007 Pharmacology 80:65-119).

The results reported in Wang et al., 2009 PLoS ONE 4(1):e4282demonstrated that acute, high-dose morphine causes an immediate buttransient switch in G protein coupling by MOR from Go to Gs similar tothe persistent switch caused by chronic morphine. Ultra-low-dose NLX orNTX prevented this switch and attenuated the chronic morphine-inducedcoupling switch by MOR. The transient nature of this acute alteredcoupling suggests the receptor eventually recovers and couples to itsnative G protein.

With chronic opioid exposure, the receptor can lose the ability torecover and continue to couple to Gs, activating the adenylylcyclase/cAMP pathway, upregulating protein kinase A, and phosphorylatingCREB as one downstream effector example. The persistently elevatedphosphorylated CREB can then shape the expression of responsive genesincluding those closely related to drug addiction and tolerance.Importantly, the equivalent blockade of Gs coupling and pS¹³³CREB by thepentapeptide binding site of naloxone (NLX) and naltrexone (NTX) on FLNAfurther elucidates the mechanism of action of ultra-low-dose NLX and NTXin their varied effects.

These data further strengthen the regulation of MOR-Gs coupling by FLNAand that binding to FLNA or using a FLNA peptide decoy for MOR canprevent the altered MOR coupling, thereby attenuating tolerance,dependence and addictive properties associated with opioid drugs.

The combination of ultra-low-dose opioid antagonists with opioidagonists formulated together in one medication has been shown toalleviate many of these undesirable aspects of opioid therapy (Burns,2005 Recent Developments in Pain Research 115-136, ISBN:81-308-0012-8).This approach shows promise for an improvement in analgesic efficacy,and animal data suggests reduced addictive potential. The identificationof the cellular target of ultra-low-dose NLX or NTX in their inhibitionof mu opioid receptor—Gs coupling as a pentapeptide segment of filamin A(Wang et al., 2008 PLoS ONE 3(2):e1554) has led to development of assaysto screen against this target to create a new generation of paintherapeutics that can provide long-lasting analgesia with minimaltolerance, dependence and addictive properties. Importantly, thenon-opioid cellular target of ultra-low-dose NLX or NTX, FLNA, providespotential for developing either a therapeutic combination of which onecomponent is not required to be ultra-low-dose, or a single-entity novelanalgesic.

Sundermann et al. U.S. Pat. No. 7,560,468 discloses a vast number ofsubstituted 1,4,9-triazaspiro[4,5]decan-2-one compounds that correspondin structure to the formula below, wherein the

various R groups can provide compounds similar to but different fromsome of those disclosed hereinafter. In particular, each of R¹ and R²can be hydrogen or any of a large number of substituents, whereas R³must be a —S(═O)₂—R⁴, —C(═S)NH—R⁵ or —C(═O)NH—R⁶; i.e., part of asulfonamide, thiourea or urea substituent. The compounds are said to beuseful for treating and/or preventing almost a complete printed patentcolumn of ailments from various types of pain, inflammation, allergies,multiple sclerosis, rheumatoid arthritis, lupus erythematosis,Tourette's syndrome, various ischemias, blood poisoning, diabetes,sepsis, cancer, particularly leukemia and/or cerebral tumor, and severalother diseases.

Published US Patent application No. 20090105290 A1 teaches substituted1-oxa-3,8-diazaspiro[4,5]decan-2-one compounds that correspond instructure to the formula below, in which the

R groups can provide a huge number of possible compounds that aresomewhat similar to, but are different from some of those disclosedhereinafter. The list of ailments for which the compounds of thisapplication are said to be useful is almost as long as that of U.S. Pat.No. 7,560,468. Included among the disease states for publishedapplication No. 20090105290 A1 are the following:pain, preferably of pain selected from the group consisting of acutepain, chronic pain, visceral pain and neuropathic pain; for theprevention and/or treatment of one or more diseases selected from thegroup consisting of disorders of food intake, preferably selected fromthe group consisting of bulimia, anorexia, obesity and cachexia; waterretention conditions; migraine; chronic paroxysmal hemicrania;depression; urinary incontinence; coughing; asthma; glaucoma; tinnitus;inflammation; neurodegenerative diseases, preferably selected from thegroup consisting of Parkinson's disease, Huntington's chorea,Alzheimer's disease and multiple sclerosis; cognitive dysfunction,preferably memory disorders; cognitive deficiency states (attentiondeficit syndrome, ADS); epilepsy; catalepsy; narcolepsy; diarrhea;gastritis; stomach ulcer; pruritus; anxiety states; panic attacks;schizophrenia; cerebral ischemic episodes; muscle spasms; cramps;gastro-esophageal reflux syndrome; alcohol and/or drug abuse, preferablynicotine and/or cocaine abuse, and/or abuse of medicines; alcohol and/ordrug dependency, preferably nicotine and/or cocaine dependency, and thelike.

The present invention identifies a compound that binds to filamin A(FLNA; the high-affinity binding site of naloxone [NLX] and naltrexone[NTX]), to reduce cell motility and inflammation as well as to preventthe Gi/o-to-Gs coupling switch of MOR and is similar to or more activethan DAMGO in activating MOR.

BRIEF SUMMARY OF THE INVENTION

The present invention contemplates an analgesic compound, a compositioncontaining that compound and a method of reducing pain in a host mammalin need thereof by administering a composition containing such acompound. A compound that corresponds in structure to Formula A iscontemplated

In Formula A, Q is CHR⁹ or C(O) [also carbonyl, or C═O], Z is CHR¹⁰ orC(O), and only one of Q and Z is C(O); each of m and n is zero or oneand the sum of m+n is 1 or 2; G, P and W are selected from the groupconsisting of NR²⁰, NR², NR⁷, S and O, where R⁷ and R² are the same ordifferent and are H, C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or3 and v+h=3, C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each ofq and r is 0, 1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbylsulfonyl or aliphatic C₁-C₁₂ hydrocarboyl, and R²⁰ is X-circle A-R¹ asdefined hereinafter, with the provisos that

i) only one of G, P and W is NR²⁰,

ii) one of G, P and W must be NR²⁰,

iii) P is NR² when other than NR²⁰;

iv) one of G and W is other than NR² or NR⁷ in which R² or R⁷ is H or analiphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) the otherof G and W is NR²⁰, NR² or NR⁷ bonded to a Z or Q, respectively, that isC(O), and

v) P is NR² in which R² is other than —S(O)₂C₁-C₃-hydrocarbyl when (a)the sum of m+n is 1 and the Q or Z that is present is CH₂, (b) the G orW that is not NR²⁰ is O, and (c) R²⁰ is —S(O)₂-phenyl-R¹, where R¹ is H,C₁-C₃-hydrocarbyl or halogen.

X is SO₂, C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O), preferably SO₂, C(O)or CH₂, and most preferably SO₂. Each of d, e, f and k is either zero orone and the sum of (d+e+f+k)=2, e is zero when d is zero, and k is zerowhen f is zero. D and F are the same or different and are CH or CD, andE and K are the same or different and are CH₂, CHD or CD₂. Each of m andn is zero or one and the sum of m+n is 1 or 2, preferably 1.

Circle A is an aromatic or heteroaromatic ring system that contains onering or two fused rings, and preferably contains a single ring. R¹represents up to three substituents, R^(1a), R^(1b), and R^(1c), thatthemselves can be the same or different, wherein each of those threegroups, R^(1a-c), is separately selected from the group consisting of H,C₁-C₆ hydrocarbyl, C₁-C₆ hydrocarbyloxy, C₁-C₆ hydrocarbyloxycarbonyl,trifluoromethyl, trifluoromethoxy, C₁-C₇ hydrocarboyl, hydroxy-,trifluoromethyl- (—CF₃) or halogen-substituted C₁-C₇ hydrocarboyl, C₁-C₆hydrocarbylsulfonyl, C₁-C₆ hydrocarbyloxysulfonyl, halogen, nitro,phenyl, cyano, carboxyl, C₁-C₇ hydrocarbyl carboxylate [C(O)O—C₁-C₇hydrocarbyl], carboxamide [C(O)NR³R⁴] or sulfonamide [S(O)₂NR³R⁴],wherein the amido nitrogen in either amide group has the formula NR³R⁴wherein R³ and R⁴ are the same or different and are H, C₁-C₄hydrocarbyl, or R³ and R⁴ together with the depicted nitrogen form a5-7-membered ring that optionally contains 1 or 2 additional heteroatoms that independently are nitrogen, oxygen or sulfur, MAr, where M is—CH₂—, —O— or —N═N— and Ar is a single-ringed aryl or heteroaryl group,and NR⁵R⁶ wherein R⁵ and R⁶ are the same or different and are H, C₁-C₄hydrocarbyl, C₁-C₄ acyl, C₁-C₄ hydrocarbylsulfonyl, or R⁵ and R⁶together with the depicted nitrogen form a 5-7-membered ring thatoptionally contains 1 or 2 additional hetero atoms that independentlyare nitrogen, oxygen or sulfur.

R⁸, R⁹, and R¹⁰ are each H, which is preferred, or two of R⁸, R⁹, andR¹⁰ are H and one is a C₁-C₈ hydrocarbyl group that is unsubstituted oris substituted with up to three atoms that are the same or different andare oxygen or nitrogen atoms.

R¹¹, R¹², R¹³ and R¹⁴ are all H, or R¹¹ and R¹³ are H and R¹² and R¹⁴are H or D, or one of the pair R¹¹ and R¹² or the pair R¹³ and R¹⁴together with the depicted ring form a saturated or unsaturated6-membered ring, and the other pair are each H or they are H and D asrecited in this paragraph (herein).

A pharmaceutically acceptable salt of a compound of Formula A and all ofthe remaining formulas disclosed herein is contemplated. Further, acontemplated compound or its pharmaceutically acceptable salt canoptionally be present in one or more forms. Illustratively, the compoundor its salt can be in the form of an individual enantiomer ordiastereoisomer. A contemplated compound or its salt can also be presentin the form of a mixture of stereoisomers. A contemplated compound orsalt can also be present in the form of a racemic mixture. Thus, acontemplated compound or its salt of Formula A can optionally be presentincluding both individual enantiomeric forms, a racemate, diastereomersand mixtures thereof.

In one preferred embodiment, a compound of Formula A corresponds instructure to Formula B.

In Formula B and the other formulas disclosed herein, substituents thathave the same designations (names) as those of Formula A have the samedefinitions, unless the formula as shown precludes part of a definitionprovided for a compound of Formula A. Thus, in a compound thatcorresponds in structure to Formula B, Q is CHR⁹ or C(O) and Z is CHR¹⁰or C(O), and only one of Q and Z is C(O). Each of m and n is zero or oneand the sum of m+n is 1 or 2. G and W are selected from the groupconsisting of NR²⁰, NR⁷, S and O, where R² and R⁷ are the same ordifferent and are H, C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or3 and v+h=3, C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each ofq and r is 0, 1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbylsulfonyl or aliphatic C₁-C₁₂ hydrocarboyl, and R²⁰ is X-circle A-R¹ asdefined hereinafter, with the provisos that

i) only one of G or W is NR²⁰,

ii) one of G and W must be NR²⁰,

iii) the G or W that is not NR²⁰ is other than NR² or NR⁷ in which R² orR⁷ is H or an aliphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and(b) the G or W that is NR²⁰ is bonded to a Z or Q, respectively, that isC(O), and

iv) R² of the depicted NR² is other than —S(O)₂C₁-C₃-hydrocarbyl when(a) the sum of m+n is 1 and the Q or Z present is CH₂, (b) the G or Wthat is not NR²⁰ is O, and (c) R²⁰ is —S(O)₂-phenyl-R¹, where R¹ is H,C₁-C₃-hydrocarbyl or halogen.

X is SO₂, C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O). Each of d, e, f andk is either zero or one and the sum of (d+e+f+k)=2, e is zero when d iszero, and k is zero when f is zero. D and F are the same or differentand are CH or CD and E and K are the same or different and are CH₂, CHDor CD₂. R² is preferably H, C(H)_(v)(D)_(h) where each of v and h is 0,1, 2 or 3 and v+h=3, C(H)_(q)(D)_(r)-aliphatic C₁-C₅ hydrocarbyl whereeach of q and r is 0, 1, or 2 and q+r=0, 1 or 2, C₁-C₆ aliphatic acyl[—C(O)—C₀-C₅ aliphatic], or a C₁-C₆ aliphatic sulfonyl [—S(O)₂—C₀-C₅aliphatic].

Circle A is an aromatic or heteroaromatic ring system that contains onering or two fused rings, preferably one ring; and group R¹ is H orrepresents up to three substituents, R^(1a), R^(1b), and R^(1c), thatthemselves can be the same or different, in which each of those threegroups, R^(1a-c), is separately selected from the group consisting of H,C₁-C₆ hydrocarbyl, C₁-C₆ hydrocarbyloxy, C₁-C₆ hydrocarbyloxycarbonyl,trifluoromethyl, trifluoromethoxy, C₁-C₇ hydrocarboyl(acyl), hydroxy-,trifluoromethyl- (—CF₃) or halogen-substituted C₁-C₇ hydrocarboyl, C₁-C₆hydrocarbylsulfonyl, C₁-C₆ hydrocarbyloxysulfonyl, halogen, nitro,phenyl, cyano, carboxyl, C₁-C₇ hydrocarbyl carboxylate, carboxamide orsulfonamide wherein the amido nitrogen in either group has the formulaNR³R⁴ wherein R³ and R⁴ are the same or different and are H, C₁-C₄hydrocarbyl, or R³ and R⁴ together with the depicted nitrogen form a5-7-membered ring that optionally contains 1 or 2 additional heteroatoms that independently are nitrogen, oxygen or sulfur, MAr, where M is—CH₂—, —O— or —N═N— and Ar is a single-ringed aryl or heteroaryl group,and NR⁵R⁶ wherein R⁵ and R⁶ are the same or different and are H, C₁-C₄hydrocarbyl, C₁-C₄ acyl, C₁-C₄ hydrocarbylsulfonyl, or R⁵ and R⁶together with the depicted nitrogen form a 5-7-membered ring thatoptionally contains 1 or 2 additional hetero atoms that independentlyare nitrogen, oxygen or sulfur.

Groups R⁸, R⁹, and R¹⁰ are each H, which is preferred, or two of R⁸, R⁹,and R¹⁰ are H and one is a C₁-C₈ hydrocarbyl group that is unsubstitutedor is substituted with up to three atoms that are the same or differentand are oxygen or nitrogen atoms.

Groups R¹¹, R¹², R¹³ and R¹⁴ are all H, or R¹¹ and R¹³ are H and R¹² andR¹⁴ are H or D, or one of the pair R¹¹ and R¹² or the pair R¹³ and R¹⁴together with the depicted ring form a saturated or unsaturated6-membered ring, and the other pair are each H or they are H and D asrecited in this paragraph (herein). A pharmaceutically acceptable saltof a compound of Formula B is also contemplated. A contemplated compoundor its salt of Formula B can optionally be present including bothindividual enantiomeric forms, a racemate, diastereomers and mixturesthereof.

In another preferred embodiment, a compound of Formula A corresponds instructure to Formula C.

In a compound that corresponds in structure to Formula C, Q is CHR⁹ orC(O) and Z is CHR¹⁰ or C(O), and only one of Q and Z is C(O). Each of mand n is zero or one and the sum of m+n is 1 or 2.

G and W are selected from the group consisting of NR², NR⁷, S and O,where R² and R⁷ are the same or different and are H, C(H)_(v)(D)_(h)where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, and X-circle A-R¹ as defined hereinafter,with the provisos that:

i) one of G and W must be NR² or NR⁷, and

ii) one of G and W is other than NR² or NR⁷ in which R² or R⁷ is H or analiphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) the otherof G and W is NR² or NR⁷ bonded to a Z or Q, respectively, that is C(O).

X is SO₂, C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O). Each of d, e, f andk is either zero or one and the sum of (d+e+f+k)=2, e is zero when d iszero, and k is zero when f is zero. D and F are the same or differentand are CH or CD and E and K are the same or different and are CH₂, CHDor CD₂. R² is preferably H, C(H)_(v)(D)_(h) where each of v and h is 0,1, 2 or 3 and v+h=3, C(H)_(q)(D)_(r)-aliphatic C₁-C₅ hydrocarbyl whereeach of q and r is 0, 1, or 2 and q+r=0, 1 or 2, C₁-C₆ aliphatic acyl[—C(O)—C₀-C₅ aliphatic], or a C₁-C₆ aliphatic sulfonyl [—S(O)₂—C₀-C₅aliphatic].

Circle A is an aromatic or heteroaromatic ring system that contains asingle ring or two fused rings; and group R¹ is H or represents up tothree substituents, R^(1a), R^(1b), and R^(1c), that themselves can bethe same or different, in which each of the three groups, R^(1a-c), isseparately selected from the group consisting of H, C₁-C₆ hydrocarbyl,C₁-C₆ hydrocarbyloxy, C₁-C₆ hydrocarbyloxycarbonyl, trifluoromethyl,trifluoromethoxy, C₁-C₇ hydrocarboyl, hydroxy-, trifluoromethyl- (—CF₃)or halogen-substituted C₁-C₇ hydrocarboyl, C₁-C₆ hydrocarbylsulfonyl,C₁-C₆ hydrocarbyloxysulfonyl, halogen, nitro, phenyl, cyano, carboxyl,C₁-C₇ hydrocarbyl carboxylate, carboxamide or sulfonamide wherein theamido nitrogen in either group has the formula NR³R⁴ wherein R³ and R⁴are the same or different and are H, C₁-C₄ hydrocarbyl, or R³ and R⁴together with the depicted nitrogen form a 5-7-membered ring thatoptionally contains 1 or 2 additional hetero atoms that independentlyare nitrogen, oxygen or sulfur, MAr (where M is —CH₂—, —O— or —N═N— andAr is a single-ringed aryl or heteroaryl group), and NR⁵R⁶ wherein R⁵and R⁶ are the same or different and are H, C₁-C₄ hydrocarbyl, C₁-C₄acyl, C₁-C₄ hydrocarbylsulfonyl, or R⁵ and R⁶ together with the depictednitrogen form a 5-7-membered ring that optionally contains 1 or 2additional hetero atoms that independently are nitrogen, oxygen orsulfur. Groups R⁸, R⁹, and R¹⁰ are each H, or two of R⁸, R⁹, and R¹⁰ areH and one is a is a C₁-C₈ hydrocarbyl group that is unsubstituted or issubstituted with up to three atoms that are the same or different andare oxygen or nitrogen atoms; and groups R¹¹, R¹², R¹³ and R¹⁴ are allH, or R¹¹ and R¹³ are H and R¹² and R¹⁴ are H or D, or one of the pairR¹¹ and R¹² or the pair R¹³ and R¹⁴ together with the depicted ring forma saturated or unsaturated 6-membered ring, and the other pair are eachH or they are H and D as recited herein (in this subparagraph). Apharmaceutically acceptable salt of a compound of Formula C is alsocontemplated. Additionally, a contemplated compound or its salt ofFormula C can optionally be present including both individualenantiomeric forms, a racemate, diastereomers and mixtures thereof.

A preferred compound of Formulas A and B has the structure of Formula I

where D and F are the same or different and are CH₂, CHD or CD₂, and W,X, Z, Q, n, m, circle A, R¹, R², R⁸ and the R groups therein defined areas described previously, except that i) when W is O, and X-circle A-R¹is —S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen, R² ofthe depicted NR² is other than —S(O)₂C₁-C₃-hydrocarbyl, and ii) W isother than NR² or NR⁷ in which R² or R⁷ is H or an aliphatic C₁hydrocarbyl when (a) the sum of m+n is 1 and (b) Z is C(O).

In another preferred embodiment, a compound of Formulas A and B has thestructure of Formula II

where D and F are the same or different and are CH₂, CHD or CD₂, andpreferably, one of D and F is CH₂; X, W, circle A, R¹, R² and the Rgroups therein defined are as described previously, except that when Wis O, and X-circle A-R¹ is —S(O)₂-phenyl-R¹, where R¹ is H,C₁-C₃-hydrocarbyl or halogen, R² of the depicted NR² is other than—S(O)₂C₁-C₃-hydrocarbyl.

In a further preferred embodiment, a compound of Formulas A and B hasthe structure of Formula III

where each of m and n is one, D and F are the same or different and areCH₂, CHD or CD₂; and W, X, Z, Q, circle A, R¹, R² and the R groupstherein defined are as described previously, except that i) one of Z andQ is C(O), and ii) W is other than NR² or NR⁷ in which R² and R⁷ is H oran aliphatic C₁ hydrocarbyl when Z is C(O).

In a still further preferred embodiment, a compound of Formulas A and Chas the structure of Formula IV

where D and F are the same or different and are CH₂, CHD or CD₂; and W,X, Z, Q, circle A, R¹, R², R⁸ and the R groups therein defined are asdescribed previously, except that i) W is other than NR² or NR⁷ in whichR² or R⁷ is H or an aliphatic C₁ hydrocarbyl when the sum of m+n is 1and Z is C(O), and ii) R² of the depicted NR² group is other than H oran aliphatic C₁ hydrocarbyl when the sum of m+n is 1, W is NR² or NR⁷,and Q is C(O).

In yet another preferred embodiment, compound of Formulas A and C hasthe structure of Formula V

where D and F are the same or different and are CH₂, CHD or CD₂; and X,W, circle A, R¹, R² and the R groups therein defined are as describedpreviously.

In still another preferred embodiment, compound of Formulas A and C hasthe structure of Formula VI

where D and F are the same or different and are CH₂, CHD or CD₂; W, X,Z, Q, circle A, R¹, R² and the R groups therein defined are as describedpreviously, and each of m and n is 1, except that i) one of Z and Q isC(O), ii) W is other than NR² or NR⁷ in which R² or R⁷ is H or analiphatic C₁ hydrocarbyl when Z is C(O), and iii) R² of the depicted NR²group is other than H or an aliphatic C₁ hydrocarbyl when W is NR² orNR⁷, and Q is C(O).

In each of the above compounds of Formulas A, B, C and I-VI, it ispreferred that X is C(O), CH₂, CD₂, or SO₂, and separately preferredthat W is NR⁷, S or O. It is also preferred that one of Q and Z be C(O)and the other be CH₂. It is further separately preferred that whereincircle A is selected from the group consisting of phenyl, pyridyl,pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl (1,3,5-triazinyl,1,2,4-triazinyl and 1,2,3-triazinyl), furanyl, thienyl, oxazolyl,isoxazolyl, thiazolyl, isothiazolyl, naphthyl, benzofuranyl,isobenzofuranyl, benzothiophenyl, isobenzothiophenyl, benzoxazolyl,benzisoxazole, quinolyl, isoquinolyl, quinazolyl, cinnolinyl,quinoxalinyl, naphthyridinyl, and benzopyrimidinyl.

As can be seen from the above definitions, a contemplated compound cancontain deuterated carbon atoms on either side of the “X” substituent.Deuterated compounds can be useful in studying the mechanism of druginteractions with living organisms for the elucidation of metabolic andbiosynthetic pathways. Deuteration can also extend the half-life of acontemplated compound in vivo because a C-D bond is stronger than a C—Hbond thereby requiring more energy input for bond cleavage. See, Blakeet al., 1975 J. Pharm. Sci. 64(3):367-391; and Nelson et al., 2003 DrugMetab. Dispos. 31(12):1481-1498, and the citations therein. Contemplateddeuterated compounds are prepared using well-known reactions.

A pharmaceutical composition is also contemplated. That compositioncomprises an above compound of Formulas A, B, C, and I-VI or itspharmaceutically acceptable salt, or an optionally present individualenantiomeric form, a racemate, a diastereomer and mixtures thereofdissolved or dispersed in a physiologically tolerable carrier. Thecompound is present in an effective analgesic amount. The composition ispreferably in solid form as in a tablet of capsule.

A method of reducing one or both of pain and inflammation in a hostmammal in need thereof is also contemplated. That method comprisesadministering to that host mammal a pharmaceutical composition asdisclosed above. The host mammal for such a method is selected from thegroup consisting of a primate, a laboratory rodent, a companion animal,and a food animal. A composition can be administered a plurality oftimes over a period of days, as well as administered a plurality oftimes in one day. That administration can be perorally or parenterally.

The present invention has several benefits and advantages.

One benefit is anti-inflammatory action combined with analgesia by acompound with a novel mechanism of action for both that does not have anarcotic structure.

An advantage of the invention is that analgesia can be provided byadministration of a contemplated composition either perorally orparenterally.

A further benefit of the invention is that as indicated by the initialdata, a contemplated compound provides the analgesic effectscharacteristic of opioid drugs but does not cause analgesic tolerance ordependence.

Another advantage of the invention as also indicated by the initial datais that a contemplated compound provides the analgesic effectscharacteristic of opioid drugs and does not have the addictive potentialof opioid drugs.

Still further benefits and advantages will be apparent to a skilledworker from the description that follows.

ABBREVIATIONS AND SHORT FORMS

The following abbreviations and short forms are used in thisspecification.

“MOR” means μ opioid receptor

“FLNA” means filamin A

“NLX” means naloxone

“NTX” means naltrexone

“Gi/o” means G protein inhibitory/other subtype, inhibits adenylylcyclase

“Gs” means G protein stimulatory subtype, stimulates adenylyl cyclase

“Gβγ” means G protein beta gamma subunit

“cAMP” means cyclic adenosine monophosphate

“CREB” means cAMP Response Element Binding protein

“IgG” means Immunoglobulin G

DEFINITIONS

In the context of the present invention and the associated claims, thefollowing terms have the following meanings:

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

As used herein, the term “hydrocarbyl” is a short hand term for anon-aromatic group that includes straight and branched chain aliphaticas well as alicyclic groups or radicals that contain only carbon andhydrogen. Inasmuch as alicyclic groups are cyclic aliphatic groups, suchsubstituents are deemed hereinafter to be subsumed within the aliphaticgroups. Thus, alkyl, alkenyl and alkynyl groups are contemplated,whereas aromatic hydrocarbons such as phenyl and naphthyl groups, whichstrictly speaking are also hydrocarbyl groups, are referred to herein asaryl groups, substituents, moieties or radicals, as discussedhereinafter. An aralkyl substituent group such as benzyl is deemed anaromatic group as being an aromatic ring bonded to an X group, where Xis CH₂. A substituent group containing both an aliphatic ring and anaromatic ring portion such as tetralin (tetrahydronaphthalene) that islinked directly through the aliphatic portion to the depicted ringcontaining the W group is deemed a non-aromatic, hydrocarbyl group. Onthe other hand, a similar group bonded directly via the aromaticportion, is deemed to be a substituted aromatic group. Where a specificaliphatic hydrocarbyl substituent group is intended, that group isrecited; i.e., C₁-C₄ alkyl, methyl or dodecenyl. Exemplary hydrocarbylgroups contain a chain of 1 to about 12 carbon atoms, and preferably 1to about 8 carbon atoms, and more preferably 1 to 6 carbon atoms of analkyl group.

A particularly preferred hydrocarbyl group is an alkyl group. As aconsequence, a generalized, but more preferred substituent can berecited by replacing the descriptor “hydrocarbyl” with “alkyl” in any ofthe substituent groups enumerated herein.

Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl,n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl,octyl, decyl, dodecyl and the like. Examples of suitable alkenylradicals include ethenyl (vinyl), 2-propenyl, 3-propenyl,1,4-pentadienyl, 1,4-butadienyl, 1-butenyl, 2-butenyl, 3-butenyl,decenyl and the like. Examples of alkynyl radicals include ethynyl,2-propynyl, 3-propynyl, decynyl, 1-butynyl, 2-butynyl, 3-butynyl, andthe like.

Usual chemical suffix nomenclature is followed when using the word“hydrocarbyl” except that the usual practice of removing the terminal“yl” and adding an appropriate suffix is not always followed because ofthe possible similarity of a resulting name to one or more substituents.Thus, a hydrocarbyl ether is referred to as a “hydrocarbyloxy” grouprather than a “hydrocarboxy” group as may possibly be more proper whenfollowing the usual rules of chemical nomenclature. Illustrativehydrocarbyloxy groups include methoxy, ethoxy, and cyclohexenyloxygroups. On the other hand, a hydrocarbyl group containing a —C(O)—functionality is referred to as a hydrocarboyl(acyl) and that containinga —C(O)O— is a hydrocarboyloxy group inasmuch as there is no ambiguity.Exemplary hydrocarboyl and hydrocarboyloxy groups include acyl andacyloxy groups, respectively, such as acetyl and acetoxy, acryloyl andacryloyloxy.

Carboxyl-related linking groups between the central spiro ring systemand an aromatic or heteroaromatic ring system, circle A, include severaltypes of ester and amide bonds. Illustrative of such bonds aresulfonamide, sulfonate and thiosulfonate esters that can be formedbetween a SO₂-containing group and an amine, oxygen or sulfur atom,respectively. Amide, ester and thioester links can be formed between anaromatic or heteroaromatic ring containing a C(O) group and a nitrogen,oxygen or sulfur atom, respectively. Similarly, a guanidino linker canbe formed between an aromatic or heteroaromatic ring containing aNHC(NH) group and a nitrogen, a urethane, carbonate or thiocarbonate canbe formed between an aromatic or heteroaromatic ring containing a OC(O)group and a nitrogen, oxygen or sulfur, respectively. A compoundcontaining a urea linker, urethane linker or isothiourea linker[NHC(O)S] can be formed between an aromatic or heteroaromatic ringcontaining a NHC(O) group and a nitrogen, oxygen or sulfur,respectively.

A “carboxyl” substituent is a —C(O)OH group. A C₁-C₆ hydrocarbylcarboxylate is a C₁-C₆ hydrocarbyl ester of a carboxyl group. Acarboxamide is a —C(O)NR³R⁴ substituent, where the R groups are definedelsewhere. Similarly, a sulfonamide is a —S(O)₂NR³R⁴ substituent, wherethe R groups are defined hereinafter. Illustrative R³ and R⁴ groups thattogether with the depicted nitrogen of a carboxamide form a 5-7-memberedring that optionally contains 1 or 2 additional hetero atoms thatindependently are nitrogen, oxygen or sulfur, include morpholinyl,piperazinyl, oxathiazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, pyrazolyl,1,2,4-oxadiazinyl and azepinyl groups.

As a skilled worker will understand, a substituent that cannot existsuch as a C₁ alkenyl or alkynyl group is not intended to be encompassedby the word “hydrocarbyl”, although such substituents with two or morecarbon atoms are intended.

The term “aryl”, alone or in combination, means a phenyl, naphthyl orother radical as recited hereinafter that optionally carries one or moresubstituents selected from hydrocarbyl, hydrocarbyloxy, halogen,hydroxy, amino, nitro and the like, such as phenyl, p-tolyl,4-methoxyphenyl, 4-(tert-butoxy)phenyl, 4-fluorophenyl, 4-chlorophenyl,4-hydroxyphenyl, and the like. The term “arylhydrocarbyl”, alone or incombination, means a hydrocarbyl radical as defined above in which onehydrogen atom is replaced by an aryl radical as defined above, such asbenzyl, 2-phenylethyl and the like. The term“arylhydrocarbyloxycarbonyl”, alone or in combination, means a radicalof the formula —C(O)—O-arylhydrocarbyl in which the term“arylhydrocarbyl” has the significance given above. An example of anarylhydrocarbyloxycarbonyl radical is benzyloxycarbonyl. The term“aryloxy” means a radical of the formula aryl-O— in which the term arylhas the significance given above. The term “aromatic ring” incombinations such as substituted-aromatic ring sulfonamide,substituted-aromatic ring sulfinamide or substituted-aromatic ringsulfenamide means aryl or heteroaryl as defined above.

As used herein, the term “binds” refers to the adherence of molecules toone another, such as, but not limited to, peptides or small moleculessuch as the compounds disclosed herein, and opioid antagonists, such asnaloxone or naltrexone.

As used herein, the term “selectively binds” refers to binding as adistinct activity. Examples of such distinct activities include theindependent binding to FLNA or a FLNA peptide, and the binding of acompound discussed above to a MOR.

As used herein, the term “FLNA-binding compound” refers to a compoundthat binds to the scaffolding protein filamin A, or more preferably to apolypeptide comprising residues -Val-Ala-Lys-Gly-Leu- (SEQ ID NO:1) ofthe FLNA sequence that correspond to amino acid residue positions2561-2565 of the FLNA protein sequence as noted in the sequence providedat the web address: UniProtKB/Swiss-Prot entry P21333, FLNA-HUMAN,Filamin-A protein sequence. A FLNA-binding compound can inhibit theMOR-Gs coupling caused by agonist stimulation of the p opioid receptorvia interactions with filamin A, preferably in the 24^(th) repeatregion. When co-administered with an opioid agonist, a FLNA-bindingcompound can enhance the analgesic effects and improve the treatment ofpain.

As used herein, the term “candidate FLNA-binding compound” refers to asubstance to be screened as a potential FLNA-binding compound. Inpreferred instances a FLNA-binding compound is also an opioid agonist.Additionally, a FLNA-binding compound can function in a combinatorymanner similar to the combination of an opioid agonist andultra-low-dose antagonist, wherein both FLNA and MOR are targeted by asingle entity.

As used herein, the term “opioid receptor” refers to a G protein coupledreceptor, located in the central nervous system that interacts withopioids. More specifically, the μ opioid receptor is activated bymorphine causing analgesia, sedation, nausea, and many other sideeffects known to one of ordinary skill in the art.

As used herein, the term “opioid agonist” refers to a substance thatupon binding to an opioid receptor can stimulate the receptor, induce Gprotein coupling and trigger a physiological response. Morespecifically, an opioid agonist is a morphine-like substance thatinteracts with MOR to produce analgesia.

As used herein, the term “opioid antagonist” refers to a substance thatupon binding to an opioid receptor inhibits the function of an opioidagonist by interfering with the binding of the opioid agonist to thereceptor.

As used herein an “analgesia effective amount” refers to an amountsufficient to provide analgesia or pain reduction to a recipient host.

As used herein an “inflammation effective amount” refers to an amountsufficient to provide reduction of inflammation to a recipient host.

As used herein the term “ultra-low-dose” or “ultra-low amount” refers toan amount of compound that when given in combination with an opioidagonist is sufficient to enhance the analgesic potency of the opioidagonist. More specifically, the ultra-low-dose of an opioid antagonistis admixed with an opioid agonist in an amount about 1000- to about10,000,000-fold less, and preferably about 10,000- to about1,000,000-fold less than the amount of opioid agonist.

As used herein an “FLNA-binding effective amount” refers to an amountsufficient to perform the functions described herein, such as reductionor prevention of inflammation, inhibition of MOR-Gs coupling, preventionof the cAMP desensitization measure, inhibition of CREB S¹³³phosphorylation and inhibition of any other cellular indices of opioidtolerance and dependence, which functions can also be ascribed toultra-low-doses of certain opioid antagonists such as naloxone ornaltrexone. When a polypeptide or FLNA-binding compound of the inventioninteracts with FLNA, an FLNA-binding effective amount can be anultra-low amount or an amount higher than an ultra-low-dose as thepolypeptide or FLNA-binding compound will not antagonize the opioidreceptor and compete with the agonist, as occurs with known opioidantagonists such as naloxone or naltrexone in amounts greater thanultra-low-doses. More preferably, when a polypeptide or VAKGL-bindingcompound of the present invention both interacts with FLNA and is anagonist of the mu opioid receptor, an FLNA-binding effective amount isan amount higher than an ultra-low-dose and is a sufficient amount toactivate the mu opioid receptor.

As used herein the phrase “determining inhibition of the interaction ofMOR with a Gs protein” refers to monitoring the cellular index of opioidtolerance and dependence caused by chronic or high-dose administrationof opioid agonists to mammalian cells. More specifically, the mu opioidreceptor—Gs coupling response can be identified by measuring thepresence of the Gus (stimulatory) subunit, the interaction of MOR withthe G protein complexes and formation of Gs-MOR coupling, theinteraction of the Gβγ protein with adenylyl cyclase types II and IV,loss of inhibition or outright enhancement of cAMP accumulation, and theactivation of CREB via phosphorylation of S¹³³.

As used herein the term “naloxone/naltrexone positive control” refers toa positive control method comprising steps discussed in a methodembodiment, wherein the candidate FLNA-binding compound is a knownopioid antagonist administered in an ultra-low amount, preferablynaloxone or naltrexone.

As used herein the term “FLNA-binding compound negative control” refersto a negative control method comprising steps discussed in a methodembodiment, wherein the candidate FLNA-binding compound is absent andthe method is carried out in the presence of only opioid agonist.

As used herein the term “pharmacophore” is not meant to imply anypharmacological activity. The term refers to chemical features and theirdistribution in three-dimensional space that constitutes and epitomizesthe preferred requirements for molecular interaction with a receptor(U.S. Pat. No. 6,034,066).

DETAILED DESCRIPTION OF THE INVENTION

It should be understood that the present disclosure is to be consideredas an exemplification of the present invention, and is not intended tolimit the invention to the specific embodiments illustrated. It shouldbe further understood that the title of this section of this application(“Detailed Description of the Invention”) relates to a requirement ofthe United States Patent Office, and should not be found to limit thesubject matter disclosed herein.

The present invention contemplates a compound that binds to FLNA andalso stimulates MOR, a composition containing that compound and methodof its use to provide one or both of pain relief and reduction ofinflammation. A contemplated compound can suppress inflammation andinhibit MOR-Gs coupling through interactions with FLNA and/or MOR.

In another aspect of the present invention, a contemplated compoundinhibits or prevents the morphine-induced Gs protein coupling by MOR.That prevention of MOR-Gs coupling is believed to occur by preserving aparticular interaction of filamin A and MOR. Downstream effects ofpreventing the MOR-Gs coupling include inhibition of cAMP accumulationand of cAMP Response Element Binding protein (CREB) activation in amanner resembling the activity of ultra-low-dose opioid antagonistsnaloxone and naltrexone.

In another aspect of the present invention, a FLNA-binding compoundprevents or inhibits the MOR-Gs coupling while itself activating MOR.

The data collected in organotypic striatal slice cultures demonstratethat after 7 days of twice daily 1-hour exposures to oxycodone, MOR instriatum switch from Go to Gs coupling (compare vehicle to oxycodoneconditions). In contrast, a compound contemplated herein does not causea switch to Gs coupling despite its ability to stimulate MOR aspreviously assessed by GTPγS binding that is blocked bybeta-funaltrexamine, a specific MOR antagonist. These data imply thatthese compounds provide the analgesic effects characteristic of opioiddrugs but do not cause analgesic tolerance or dependence, and do nothave the addictive potential of opioid drugs.

A compound contemplated by the present invention binds to anabove-defined FLNA polypeptide as well as stimulates MOR. A contemplatedcompound corresponds in structure to Formula A

In Formula A,

Q is CHR⁹ or C(O), Z is CHR¹⁰ or C(O), and only one of g and Z is C(O);each of m and n is zero or one and the sum of m+n is 1 or 2, preferably1; each of G, P and W is selected from the group consisting of NR²⁰,NR², NR⁷, S and O, where R⁷ and R² are the same or different and are H,C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl, aliphaticC₁-C₁₂ hydrocarbyl sulfonyl or aliphatic C₁-C₁₂ hydrocarboyl(acyl), andR²⁰ is X-circle A-R¹ as defined hereinafter, with the provisos that:

i) only one of G, P and W is NR²⁰,

ii) one of G, P and W must be NR²⁰,

iii) P is NR² when other than NR²⁰,

iv) one of G and W is other than NR² or NR⁷ in which R² and R⁷ is H oran aliphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) theother of G and W is NR²⁰, NR², or NR⁷ bonded to a Z or Q, respectively,that is C(O), and

v) P is NR² in which R² is other than —S(O)₂C₁-C₃-hydrocarbyl when (a)the sum of m+n is 1 and the Q or Z present is CH₂, (b) the G or W thatis not NR²⁰ is O, and (c) R²⁰ is —S(O)₂-phenyl-R¹, where R¹ is H,C₁-C₃-hydrocarbyl or halogen.

X is SO₂, C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O), preferably SO₂, C(O)or CH₂, and most preferably SO₂.

Each of d, e, f and k is either zero or one and the sum of (d+e+f+k)=2,e is zero when d is zero, and k is zero when f is zero; D and F are thesame or different and are CH or CD; and E and K are the same ordifferent and are CH₂, CHD or CD₂.

Circle A is an aromatic or heteroaromatic ring system that preferablycontains a single ring, but can also contain two fused rings. R¹ is H orrepresents up to three substituents, R^(1a), R^(1b), and R^(1c), thatthemselves can be the same or different, wherein each of those threegroups, R^(1a-c), is separately selected from the group consisting of H,C₁-C₆ hydrocarbyl, C₁-C₆ hydrocarbyloxy, C₁-C₆ hydrocarbyloxycarbonyl,trifluoromethyl, trifluoromethoxy, C₁-C₇ hydrocarboyl, hydroxy-,trifluoromethyl- (—CF₃) or halogen-substituted C₁-C₇ hydrocarboyl, C₁-C₆hydrocarbylsulfonyl, C₁-C₆ hydrocarbyloxysulfonyl, halogen, nitro,phenyl, cyano, carboxyl, C₁-C₇ hydrocarbyl carboxylate [C(O)O—C₁-C₇hydrocarbyl], carboxamide [C(O)NR³R⁴] or sulfonamide [S(O)₂NR³R⁴],wherein the amido nitrogen in either amide group has the formula NR³R⁴in which R³ and R⁴ are the same or different and are H, C₁-C₄hydrocarbyl, or R³ and R⁴ together with the depicted nitrogen form a5-7-membered ring that optionally contains 1 or 2 additional heteroatoms that independently are nitrogen, oxygen or sulfur, MAr, where M is—CH₂—, —O— or —N═N— and Ar is a single-ringed aryl or heteroaryl groupand NR⁵R⁶ wherein R⁵ and R⁶ are the same or different and are H, C₁-C₄hydrocarbyl, C₁-C₄ acyl, C₁-C₄ hydrocarbylsulfonyl, or R⁵ and R⁶together with the depicted nitrogen form a 5-7-membered ring thatoptionally contains 1 or 2 additional hetero atoms that independentlyare nitrogen, oxygen or sulfur.

R⁸, R⁹, and R¹⁰ are each H, which is preferred, or two of R⁸, R⁹, andR¹⁰ are H and one is a C₁-C₈ hydrocarbyl group that is unsubstituted oris substituted with up to three atoms that are the same or different andare oxygen or nitrogen atoms.

R¹¹, R¹², R¹³ and R¹⁴ are all H, or R¹¹ and R¹³ are H and R¹² and R¹⁴are H or D, or one of the pair R¹¹ and R¹² or the pair R¹³ and R¹⁴together with the depicted ring form a saturated or unsaturated6-membered ring, and the other pair are each H or they are H and D asrecited herein (in this subparagraph).

A pharmaceutically acceptable salt of a compound of Formula A and all ofthe remaining formulas disclosed herein is also contemplated. A compoundhaving an asymmetrical (chiral) carbon or a salt thereof can exist inthe form of two enantiomers. The invention relates both to eachenantiomer and to their mixture; i.e., to both enantiomeric forms and totheir mixture. Additionally, where two or more chiral centers arepresent, diastereomers can form.

Where a contemplated compound or a pharmaceutically acceptable salt ofFormula A or any of the other formulas herein is obtained in the form ofa mixture of the stereoisomers, preferably in the form of the racematesor other mixtures of the various enantiomers and/or diastereoisomers,they can be separated and optionally isolated by conventional methodsknown to the person skilled in the art. Illustratively, for example,chromatographic separation processes are useful, particularly liquidchromatography processes under standard pressure or under elevatedpressure, preferably MPLC and HPLC methods, and also methods involvingfractional crystallization. This can particularly involve the separationof individual enantiomers, e.g., diastereoisomeric salts separated bymeans of HPLC in the chiral phase or by means of crystallization withchiral acids, for example (+)-tartaric acid, (−)-tartaric acid, or(+)-10-camphorsulfonic acid. An enantiomer separated by chiral saltformation can readily be converted into an achiral or racemicpharmaceutically acceptable salt for use.

A compound of Formula A or a pharmaceutically acceptable salt thereof iscontemplated to be optionally present in enantiomerically pure form;i.e., in (S) or (R) configuration or d and l forms, or in the form of aracemic mixture showing an (S,R) or (d,l) configuration, or as one ormore diastereomers, and mixtures thereof. Similarly, a compound ofFormula A or its pharmaceutically acceptable salt can optionally be usedin a process of the invention in enantiomerically pure form; i.e., in(S) or (R) configuration or d and l forms, or in the form of a racemicmixture showing an (S,R) or (d,l) configuration, or as one or morediastereomers, and mixtures thereof.

Thus, a contemplated compound or its pharmaceutically acceptable saltcan optionally be present in one or more forms. Illustratively, thecompound or its salt can be in the form of an individual enantiomer ordiastereoisomer. A contemplated compound or its salt can also be presentin the form of a mixture of stereoisomers. A contemplated compound orsalt can also be present in the form of a racemic mixture.

In preferred embodiments, a compound of Formula A corresponds instructure to either Formula B or Formula C, and can be present as apharmaceutically acceptable salt, and can optionally be presentincluding both individual enantiomeric forms, a racemate, diastereomersand mixtures thereof.

In a compound that corresponds in structure to Formula B, G and W areselected from the group consisting of NR²⁰, NR⁷, S and O, where R² andR⁷ are the same or different and are C(H)_(v)(D)_(h) where each of v andh is 0, 1, 2 or 3 and v+h=3, C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁hydrocarbyl where each of q and r is 0, 1, or 2 and q+r=0, 1 or 2,aliphatic C₁-C₁₂ hydrocarbyl sulfonyl or aliphatic C₁-C₁₂ hydrocarboyl,and R²⁰ is X-circle A-R¹, with the provisos that:

i) only one of G and W is NR²⁰,

ii) one of G and W must be NR²⁰,

iii) the G or W that is not NR²⁰ is other than NR² or NR⁷ in which R² orR⁷ is H or an aliphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and(b) the G or W that is NR²⁰ is bonded to a Z or Q, respectively, that isC(O), and

iv) R² of the depicted NR² is other than —S(O)₂C₁-C₃-hydrocarbyl when(a) the sum of m+n is 1 and the Q or Z that is present is CH₂, (b) the Gor W that is not NR²⁰ is O, and (c) R²⁰ is —S(O)₂-phenyl-R¹, where R¹ isH, C₁-C₃-hydrocarbyl or halogen.

For Formula C, G and W are selected from the group consisting of NR²,NR⁷, S and O, where R² and R⁷ are the same or different and are H,C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, with the provisos that:

i) one of G and W must be NR² or NR⁷, and

ii) one of G and W is other than NR² or NR⁷ in which R² or R⁷ is H or analiphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) the otherof G and W is NR² or NR⁷ bonded to a Z or Q, respectively, that is C(O).

In both of Formulas B and C, the symbols X, Z, Q, d, e, f, g, n, m,circle A, and all of the R groups not otherwise defined in theparagraphs below their structural formulas are as defined previously.

A preferred compound of one or more of Formulas A, B and C includes thefollowing:

In one embodiment, a preferred compound of Formulas A and B has thestructure of Formula I

wherein D and F are the same or different and are CH₂, CHD or CD₂; andW, X, Z, Q, n, m, circle A, R¹, R², R⁸ and the R groups therein definedare as described previously, except that i) R² of the depicted NR² isother than —S(O)₂C₁-C₃-hydrocarbyl when (a) the sum of m+n is 1 and theQ or Z present is CH₂, (b) the G or W that is not NR²⁰ is O, and (c) R²⁰is —S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen, andii) W is other than NR² or NR⁷ in which R² or R⁷ is H or an aliphatic C₁hydrocarbyl when (a) the sum of m+n is 1 and (b) Z is C(O).

In another preferred embodiment where R⁸ is H, one of n and m is zeroand the remaining Z or Q is CH₂, a compound of Formulas A, B and I hasthe structure of Formula II

wherein D and F are the same or different and are CH₂, CHD or CD₂; andX, W, circle A, R¹, R² and the R groups therein defined are as describedpreviously, except that R² of the depicted NR² is other than—S(O)₂C₁-C₃-hydrocarbyl when W is O, and X-circle A-R¹ is—S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen.

In a further preferred embodiment, where R⁸ is H, a compound of FormulasA, B and I has the structure of Formula III

wherein D and F are the same or different and are CH₂, CHD or CD₂; eachof m and n is one; and W, X, Z, Q, circle A, R¹, R² and the R groupstherein defined are as described previously, except that i) one of Z andQ is C(O), and ii) W is other than NR² or NR⁷ in which R² and R⁷ is H oran aliphatic C₁ hydrocarbyl when Z is C(O).

In a still further preferred embodiment, a compound of Formulas A and Chas the structure of Formula IV

wherein D and F are the same or different and are CH₂, CHD or CD₂; andW, X, Z, Q, circle A, R¹, R², R⁸ and the R groups therein defined are asdescribed previously, except that i) W is other than NR² or NR⁷ in whichR² or R⁷ is H or an aliphatic C₁ hydrocarbyl when the sum of m+n is 1and Z is C(O), and ii) R² of the depicted NR² group is other than H oran aliphatic C₁ hydrocarbyl when the sum of m+n is 1, W is NR² or NR⁷,and Q is C(O).

In yet another preferred embodiment where R⁸ is H, one of n and m iszero and the remaining Z or Q is CH₂, a compound of Formulas A, C and IVhas the structure of Formula V

wherein D and F are the same or different and are CH₂, CHD or CD₂; andX, W, circle A, R¹, R² and the R groups therein defined are as describedpreviously.

In still another preferred embodiment, where R⁸ is H, a compound ofFormulas A, C and I has the structure of Formula VI

wherein D and F are the same or different and are CH₂, CHD or CD₂; andeach of m and n is one; W, X, Z, Q, circle A, R¹, R² and the R groupstherein defined are as described previously, and each of m and n is 1,except that i) one of Z and Q is C(O), ii) W is other than NR² or NR⁷ inwhich R² or R⁷ is H or an aliphatic C₁ hydrocarbyl when Z is C(O), andiii) R² of the depicted NR² group is other than H or an aliphatic C₁hydrocarbyl when W is NR² or NR⁷, and Q is C(O).

Again, a pharmaceutically acceptable salt of all of the compounds ofFormulas A, B, C, and I-VI is contemplated. It is also noted that thepreviously mentioned preferences regarding apply to X, W, Z, Q, d, e, f,k, n, m, circle A, and all of the R groups apply to a compound ofFormulas A, B, C, and I-VI.

In a compound of Formulas A, B and I, II and III, for enhanced potencyas a MOR agonist, it is preferred that R¹ have a Hammett sigma value fora para-position substituent that is about −0.7 to about zero, and morepreferably, a Hammett sigma value for a para-position substituent thatis about −0.3 to about −0.1. An R² group is preferably a H or C₁-C₆hydrocarbyl, with a cyclopropylmethyl group being preferred.

In a compound of Formulas A, C and IV, V and VI, for enhanced potency asa MOR agonist, R¹ preferably has a Hammett sigma value for apara-position substituent that is greater than −0.2, and morepreferably, a Hammett sigma value for a para-position substituent thatis zero or positive (greater than zero). For enhanced specificity, it isalso preferred that G and W are both NR² or NR⁷ and that R² and R⁷ bothbe H, so that both G and W are NH, and that one of Z and Q being C(O).

Hammett sigma values are well known in organic chemistry and thosevalues for para-position substituents reflect both electron donation orwithdrawal via an inductive effect, but also are understood to reflect aresonance effect. It is noted that the recited para-position sigma valueis utilized regardless of the actual position of the substituent on thearomatic ring. For Hammett sigma values see, for example, U.S. Pat. No.7,473,477, No. 5,811,521, No. 4,746,651, and No. 4,548,905. A list ofHammett sigma values can be found in J. Hine, Physical OrganicChemistry, 2^(nd) ed., McGraw-Hill Book Co., Inc., New York page 87(1962) and at the web site: wiredchemist.com/chemistry/data/hammettsigma constants.

A contemplated aromatic ring (aryl) system of circle A of one of thecontemplated compounds preferably contains a single aromatic ring, butcan also contain two fused aromatic rings. An illustrative circle Aaromatic ring system is selected from the group consisting of phenyl,pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl(1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl), furanyl,thienyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, naphthyl,benzofuranyl, isobenzofuranyl, benzothiophenyl, isobenzothiophenyl,benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl, quinazolyl,cinnolinyl, quinoxalinyl, naphthyridinyl, and benzopyrimidinyl.

An illustrative single-ringed aryl or heteroaryl group of a circle Agroup or of a substituent of circle A, MAr, is selected from the groupconsisting of phenyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl,triazinyl (1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl),furanyl, thienyl, oxazolyl, isoxazolyl, thiazolyl and isothiazolylgroups.

Phenyl, pyridinyl and furanyl are a preferred aromatic or heteroaromaticring system of circle A, with phenyl being more preferred. Phenyl,pyridinyl and furanyl are also preferred single-ringed aryl orheteroaryl groups, Ar, of a MAr substituent, with phenyl beingparticularly preferred.

X and Y can form a sulfonamido, a carboxamido, a urea, a guanidino ormethylene linkage from the circle A ring system to a depicted nitrogenatom of the central spiro rings.

Examining a compound of the above formulas more closely, it is seen thatthat formula defines a double ringed, substituted spiro compound thatcan have two six-membered rings or one six- and one five-membered ring,as when one of “m” and “n” is one and the other zero. One of those rings(the lower ring in the formulas) contains one nitrogen atom in the6-membered ring and the remaining ring atoms are carbons. The ring thatcan contain 5- or 6-ring atoms (upper ring in the formulas) can containone ring nitrogen and four or five carbons, or two nitrogens, a nitrogenand a sulfur or a nitrogen and an oxygen atom along with three or fourring carbons. Illustrative central spiro rings are shown below wherewavy lines are used to indicate the presence of covalent bonds to otherentities, and where R⁷ is defined above and R⁸ is H for clarity.

Illustrative compounds of Formula A in which d and e are each zero andR¹¹, R¹² and R¹³ are each H have asymmetric spiro ring structures a fewof which are shown below with wavy lines indicating the presence ofcovalent bonds to other entities, and R⁷ is defined above and R⁸ isagain H for clarity.

In preferred practice for the compounds of Formulas A, B and C, n iszero, e and g are both zero and R¹¹, R¹² and R¹³ are all H, so thecentral ring is a spiro 5,6-ring system whose 6-membered ring isunsubstituted and in which the spiro bonds are in the 4-positionrelative to the nitrogen of the 6-membered ring. It is separatelypreferred that W be O, S or NR⁷. It is also preferred that X be SO₂(sulfonyl).

The aromatic substituent, the circle A, is linked to one nitrogen atomof the spiro rings by a X group that is SO₂, C(O), CH₂, CD₂, OC(O),NHC(NH) or NHC(O), preferably SO₂, C(O), CH₂, or CD₂, and mostpreferably SO₂. The resulting aromatic substituent is thereby linked tothe spiro ring portion by a sulfonamide, an amide, a methylene, a ureaor a urethane linkage. Aryl sulfonamide bridges, aryl amide bridges andphenylmethylene bridges (benzyl compounds) are preferred, with arylsulfonamides being particularly preferred.

A particularly preferred compound of Formulas A and B, and that embodiesone or more of the above separate preferences is a compound whosestructure corresponds to that of Formula I

A particularly preferred compound of Formula I is selected from thegroup consisting of

In another embodiment, a contemplated compound corresponds in structureto Formula II

Here, particularly preferred compounds include those selected from thegroup consisting of

In a further embodiment, a particularly preferred compound of Formulas Aand B is a compound of Formula III

A particular compound of Formula III is selected from the groupconsisting of

In a further embodiment, a particularly preferred compound of Formulas Aand C is a compound of Formula IV

A particularly preferred compound of Formula IV has the structure

In yet another embodiment, a particularly preferred compound of FormulasA and C is a compound of Formula V

A particularly preferred compound of Formula V corresponds in structureto

In still another preferred embodiment, compound of Formulas A and C hasthe structure of Formula VI

A particularly preferred compound of Formula VI corresponds in structureto

It is to be understood that before-discussed Formula A is intended to begeneric and encompass each of Formulas B and C, and their compounds.Formula B and Formula C are intended to define non-overlapping groups ofcompounds. On the other hand, Formulas I-III define partiallyoverlapping groups of compounds, all of which are encompassed by FormulaB. Similarly, Formulas IV-VI define another partially overlapping groupof compounds that are encompassed by Formula C. Thus, Formulas II andIII define two non-overlapping groups of compounds that are among thosedefined by Formula I. Similarly, Formulas V and VI define twonon-overlapping groups of compounds that are among those defined byFormula IV.

The present invention also contemplates a method of treatment to reduceone or both of pain and inflammation in a treated mammal. A compound ofFormulas A, B, C, I, II, III, IV, V and VI or its pharmaceuticallyacceptable salt present in an analgesic effective amount dissolved ordispersed in a physiologically tolerable diluent can be and preferablyis used in such a treatment. Such compositions and methods are discussedfurther hereinafter.

In another aspect, a contemplated compound is selected in part using amethod for determining the ability of a candidate FLNA-binding compound,other than naloxone or naltrexone, to inhibit the interaction of the muopioid receptor with filamin A (FLNA) and thereby prevent the mu opioidreceptor from coupling to Gs proteins (Gs). That method comprises thesteps of: (a) admixing the candidate FLNA-binding compound (alone ifsuch FLNA-binding compound also stimulates MOR or with a MOR agonistotherwise) with mammalian cells that contain the mu opioid receptor andFLNA in their native conformations and relative orientations, the opioidagonist being present in an agonist effective amount and/or beingadministered in a repeated, chronic manner the FLNA-binding compoundbeing present in an FLNA-binding effective amount; and (b) determininginhibition of the interaction of the mu opioid receptor with the Gprotein by analysis of the presence or the absence of the Gαs subunit ofGs protein, wherein the absence of the Gαs subunit indicates inhibitionof the interaction of the mu opioid receptor with the Gs protein.

In one aspect, the analysis of Gs protein coupling by the mu opioidreceptor and downstream effects elicited by admixing mammalian cellswith a before-defined compound can be conducted by any one or more ofseveral methods such as for example co-immunoprecipitation of Gaproteins with MOR, Western blot detection of MOR in immunoprecipitates,and densitometric quantification of Western blots.

Pharmaceutical Composition

A compound of the invention can be provided for use by itself, or as apharmaceutically acceptable salt. Although substituent groups canprovide an acid functionality, a contemplated compound of any ofFormulas A-C and Formulas I-VI is an amine and can typically be used inthe form of a pharmaceutically acceptable acid addition salt derivedfrom an inorganic or organic acid. Exemplary salts include but are notlimited to the following: acetate, adipate, alginate, citrate,aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate,camphorsulfonate, digluconate, cyclopentanepropionate, dodecylsulfate,ethanesulfonate, glucoheptanoate, glycerophosphate, hemisulfate,heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide,hydroiodide, 2-hydroxy-ethanesulfonate, lactate, maleate,methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, palmoate,pectinate, persulfate, 3-phenylpropionate, picrate, pivalate,propionate, succinate, tartrate, thiocyanate, tosylate, mesylate andundecanoate.

Other compounds useful in this invention that contain acidfunctionalities can also form salts. Examples include salts with alkalimetals or alkaline earth metals, such as sodium, potassium, calcium ormagnesium or with organic bases or basic quaternary ammonium salts.

The reader is directed to Berge, 1977 J. Pharm. Sci. 68(1):1-19 forlists of commonly used pharmaceutically acceptable acids and bases thatform pharmaceutically acceptable salts with pharmaceutical compounds.

In some cases, the salts can also be used as an aid in the isolation,purification or resolution of the compounds of this invention. In suchuses, the acid used and the salt prepared need not be pharmaceuticallyacceptable.

A contemplated composition can be used in the manufacture of amedicament that is useful at least for lessening or reducing pain in amammal that is in need, such as somatic, visceral, neuropathic orsympathetic pain, including musculoskeletal pain, inflammatory pain,burn pain, and pain from syndromes such as fibromyalgia and complexregional pain syndrome (CRPS). A contemplated composition can also beused in the manufacture of a medicament that is useful in reducinginflammation. Inasmuch as pain and inflammation are not alwayscoincident, a contemplated composition is referred to as being used toreduce one or both of pain and inflammation, or a similar phrase.

A contemplated pharmaceutical composition contains an analgesiaeffective amount of a compound Formulas A, B or C and of Formulas I-VIor a pharmaceutically acceptable salt thereof dissolved or dispersed ina physiologically tolerable carrier. Such a composition can beadministered to mammalian cells in vitro as in a cell culture, or invivo as in a living, host mammal in need.

A contemplated composition is typically administered a plurality oftimes over a period of days. More usually, a contemplated composition isadministered a plurality of times in one day.

As is seen from the data that follow in the table below, a contemplatedcompound is active in the assays studied at micromolar amounts. In thelaboratory mouse tail flick test, orally administered morphine exhibitedan A₅₀ value of 61.8 (52.4-72.9) mg/kg, and a mean maximumantinociception amount of about 43% at 56 mg/kg at about 20 minutes.Several orally administered compounds whose structures are shown in theTable of Correspondence hereinafter were compared at various dosages todetermine a maximal amount of pain relief (antinociception amount) andthe time after administration that that maximal pain relief occurred.Data from those studies are shown below.

Maximum Compound Antinociception Dosage Time to Max. Number (%) (mg/kg)(minutes) Morphine 43 56 20 C0027 70 56 20 C0066 55 56 20 C0134M 50 5630 C0108M 50 56 20 C0090 35 56 20 C0089 30 56 20 S-C0027 90 100 30S-C0027 70 56 30 F-C0134 93 56 20It is thus seen that the contemplated compounds are quite active andpotent, and that a skilled worker can readily determine an appropriatedosage level to achieve a desired amount of pain reduction, particularlyin view of the relative activity of a contemplated compound compared toorally administered morphine.

A contemplated pharmaceutical composition can be administered orally(perorally), parenterally, by inhalation spray in a formulationcontaining conventional nontoxic pharmaceutically acceptable carriers,adjuvants, and vehicles as desired. The term parenteral as used hereinincludes subcutaneous injections, intravenous, intramuscular,intrasternal injection, or infusion techniques. Formulation of drugs isdiscussed in, for example, Hoover, John E., Remington's PharmaceuticalSciences, Mack Publishing Co., Easton, Pa.; 1975 and Liberman, H. A. andLachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York,N.Y., 1980.

Injectable preparations, for example, sterile injectable aqueous oroleaginous suspensions can be formulated according to the known artusing suitable dispersing or wetting agents and suspending agents. Thesterile injectable preparation can also be a sterile injectable solutionor suspension in a nontoxic parenterally acceptable diluent or solvent,for example, as a solution in 1,3-butanediol. Among the acceptablevehicles and solvents that can be employed are water, Ringer's solution,and isotonic sodium chloride solution, phosphate-buffered saline. Liquidpharmaceutical compositions include, for example, solutions suitable forparenteral administration. Sterile water solutions of an activecomponent or sterile solution of the active component in solventscomprising water, ethanol, or propylene glycol are examples of liquidcompositions suitable for parenteral administration.

In addition, sterile, fixed oils are conventionally employed as asolvent or suspending medium. For this purpose any bland fixed oil canbe employed including synthetic mono- or diglycerides. In addition,fatty acids such as oleic acid find use in the preparation ofinjectables. Dimethyl acetamide, surfactants including ionic andnon-ionic detergents, polyethylene glycols can be used. Mixtures ofsolvents and wetting agents such as those discussed above are alsouseful.

Sterile solutions can be prepared by dissolving the active component inthe desired solvent system, and then passing the resulting solutionthrough a membrane filter to sterilize it or, alternatively, bydissolving the sterile compound in a previously sterilized solvent understerile conditions.

Solid dosage forms for oral administration can include capsules,tablets, pills, powders, and granules. In such solid dosage forms, thecompounds of this invention are ordinarily combined with one or moreadjuvants appropriate to the indicated route of administration. Ifadministered per os, the compounds can be admixed with lactose, sucrose,starch powder, cellulose esters of alkanoic acids, cellulose alkylesters, talc, stearic acid, magnesium stearate, magnesium oxide, sodiumand calcium salts of phosphoric and sulfuric acids, gelatin, acacia gum,sodium alginate, polyvinylpyrrolidone, and/or polyvinyl alcohol, andthen tableted or encapsulated for convenient administration. Suchcapsules or tablets can contain a controlled-release formulation as canbe provided in a dispersion of active compound in hydroxypropylmethylcellulose. In the case of capsules, tablets, and pills, the dosage formscan also comprise buffering agents such as sodium citrate, magnesium orcalcium carbonate or bicarbonate. Tablets and pills can additionally beprepared with enteric coatings.

A mammal in need of treatment and to which a pharmaceutical compositioncontaining a contemplated compound is administered can be a primate suchas a human, an ape such as a chimpanzee or gorilla, a monkey such as acynomolgus monkey or a macaque, a laboratory animal such as a rat, mouseor rabbit, a companion animal such as a dog, cat, horse, or a foodanimal such as a cow or steer, sheep, lamb, pig, goat, llama or thelike.

Where in vitro mammalian cell contact is contemplated, a CNS tissueculture of cells from an illustrative mammal is often utilized, as isillustrated hereinafter. In addition, a non-CNS tissue preparation thatcontains opioid receptors such as guinea pig ileum can also be used.

Preferably, the pharmaceutical composition is in unit dosage form. Insuch form, the composition is divided into unit doses containingappropriate quantities of the active urea. The unit dosage form can be apackaged preparation, the package containing discrete quantities of thepreparation, for example, in vials or ampules.

EXAMPLES

The present invention is described in the following examples which areset forth to aid in the understanding of the invention, and should notbe construed to limit in any way the invention as defined in the claimswhich follow thereafter.

The experiments described herein were carried out on organotypicstriatal slices from male Sprague Dawley rats (200 to 250 g) purchasedfrom Taconic (Germantown, N.Y.). Rats were housed two per cage andmaintained on a regular 12-hour light/dark cycle in a climate-controlledroom with food and water available ad libitum and sacrificed by rapiddecapitation. All data are presented as mean±standard error of the mean.Treatment effects were evaluated by two-way ANOVA followed byNewman-Keul's test for multiple comparisons. Two-tailed Student's t testwas used for post hoc pairwise comparisons. The threshold forsignificance was p<0.05.

The following Table of Correspondence shows the structures of thecompounds discussed herein and their identifying numbers.

Tables A-AE hereinafter illustrate several further contemplatedcompounds having various linking groups X, central spiro ring systemsand aromatic ring systems, circle A, and wherein the wavy lines indicatethe place of bonding between the circle A ring system and the centralspiro ring system. Substituents on the aromatic or heteroaromatic ringsystems are omitted for added clarity with the understanding that one tothree substituents, R^(1a-c), can be present bonded to each of the ringsystems as discussed previously.

TABLE A

TABLE B

  R¹ = H

TABLE C

  R¹ = H

TABLE D

  R¹ = H

TABLE E

  R¹ = H

TABLE F

  R¹ = H

TABLE G

  R¹ = H

TABLE H

  R¹ = H

TABLE I

  R¹ = H

TABLE J

  R¹ = H

TABLE K

  R¹ = H

TABLE L

  R¹ = H

TABLE M

  R¹ = H

TABLE N

  R¹ = H

TABLE O

  R¹ = H

TABLE P

  R¹ = H

TABLE Q

TABLE R

TABLE S

TABLE T

TABLE U

TABLE V

TABLE W

TABLE X

TABLE Y

TABLE Z

TABLE AA

TABLE AB

TABLE AC

TABLE AD

TABLE AE

Without departing from the spirit and scope of this invention, one ofordinary skill can make various changes and modifications to theinvention to adapt it to various usages and conditions. As such, thesechanges and modifications are properly, equitably, and intended to be,within the full range of equivalence of the following claims.

Example 1 MOR Agonist Activity using GTPγS Binding Assay

To assess the mu opiate receptor (MOR) agonist activity of positivecompounds from the FLNA screening, compounds were tested in a [³⁵S]GTPγSbinding assay using striatal membranes. Our previous study has shownthat in striatal membranes, activation of MOR leads to an increase in[³⁵S]GTPγS binding to Gαo (Wang et al., 2005 Neuroscience 135:247-261).

Striatal tissue was homogenized in 10 volumes of ice cold 25 mM HEPESbuffer, pH 7.4, which contained 1 mM EGTA, 100 mM sucrose, 50 μg/mlleupeptin, 0.04 mM PMSF, 2 μg/ml soybean trypsin inhibitor and 0.2%2-mercaptoethanol. The homogenates were centrifuged at 800×g for 5minutes and the supernatants were centrifuged at 49,000×g for 20minutes. The resulting pellets were suspended in 10 volume of reactionbuffer, which contained 25 mM HEPES, pH 7.5, 100 mM NaCl, 50 μg/mlleupeptin, 2 μg/ml soybean trypsin inhibitor, 0.04 mM PMSF and 0.02%2-mercaptomethanol.

The resultant striatal membrane preparation (200 μg) was admixed andmaintained (incubated) at 30° C. for 5 minutes in reaction buffer asabove that additionally contained 1 mM MgCl₂ and 0.5 nM [³⁵S]GTPγS (0.1μCi/assay, PerkinElmer Life and Analytical Sciences) in a total volumeof 250 μl and continued for 5 minutes in the absence or presence of0.1-10 μM of an assayed compound of interest. The reaction wasterminated by dilution with 750 μl of ice-cold reaction buffer thatcontained 20 mM MgCl₂ and 1 mM EGTA and immediate centrifugation at16,000×g for 5 minutes.

The resulting pellet was solubilized by sonicating for 10 seconds in 0.5ml of immunoprecipitation buffer containing 0.5% digitonin, 0.2% sodiumcholate and 0.5% NP-40. Normal rabbit serum (1 μl) was added to 1 ml oflysate and incubated at 25° C. for 30 minutes. Nonspecific immunecomplexes were removed by incubation with 25 μl of proteinA/G-conjugated agarose beads at 25° C. for 30 minutes followed bycentrifugation at 5,000×g at 4° C. for 5 minutes. The supernatant wasdivided and separately incubated at 25° C. for 30 minutes withantibodies raised against Gαo proteins (1:1,000 dilutions).

The immunocomplexes so formed were collected by incubation at 25° C. for30 minutes with 40 μl of agarose-conjugated protein A/G beads andcentrifugation at 5,000×g at 4° C. for 5 minutes. The pellet was washedand suspended in buffer containing 50 mM Tris-HCl, pH 8.0, and 1% NP-40.The radioactivity in the suspension was determined by liquidscintillation spectrometry. The specificity of MOR activation of[³⁵S]GTPγS binding to Gαo induced by a selective compound was defined byinclusion of 1 μM β-funaltrexamine (β-FNA; an alkylating derivative ofnaltrexone that is a selective MOR antagonist). DAMGO(H-Tyr-D-Ala-Gly-N-MePhe-Gly-OH; 1 or 10 μM) was used as a positivecontrol.

The results of this study are shown in the Table below.

FLNA-Binding Compound MOR Agonist Activity FLNA- Concentration ofFLNA-Binding Compound as Agonist Binding 1 μM + % DAMGO % DAMGO %DAMGO + Compound 0.1 μM 1 μM BFNA (0.1 μM) (1 μM) BFNA C0011 217.0%305.0% 19.0% 126.8% 114.3% 36.5% C0026 207.2% 288.4% 21.2% 107.7% 105.6%48.3% C0027 233.2% 313.9% 72.2% 121.3% 115.0% 164.5% S-C0027 156.2%286.8% 56.2% 74.2% 84.4% 98.1% C0034-3 — — — — — — C0037-2 — — — — — —C0040 145.8% 308.3% 90.4% 93.1% 133.2% 277.3% C0043 175.4% 242.6% 83.3%103.8% 110.9% 501.8% C0044 173.7% 280.1% 59.1% 102.8% 128.0% 356.0%C0045 149.2% 238.8% 105.3% 88.3% 109.1% 634.3% C0046 286.2% 492.9%156.8% 197.4% 211.5% 356.4% C0050 110.3% 127.6% 59.0% 76.1% 54.8% 134.1%C0055 — — — — — — C0056 98.6% 193.4% 86.3% 68.0% 83.0% 196.1% C0060166.5% 218.9% 143.9% 114.8% 93.9% 327.0% C0086M 206.8% 265.3% 152.3%117.5% 104.9% 692.3% C0087M 262.8% 329.6% 142.5% 138.9% 132.8% 293.8%C0088M 276.3% 355.3% 177.1% 160.5% 161.7% 513.3% C0089M 234.5% 295.3%81.9% 136.3% 134.4% 237.4% C0090M 237.0% 341.0% 41.0% 137.7% 155.2%118.8% C0091M 207.9% 274.4% 80.8% 118.1% 108.5% 367.3% C0093M 140.0%211.8% 44.0% 81.3% 96.4% 127.5% C0094M 172.5% 263.5% 115.3% 100.2%119.9% 334.2% C0095M 189.1% 224.6% 107.7% 107.4% 88.8% 489.5% C0096M186.4% 328.9% 127.1% 105.9% 130.0% 577.7% C0099M 157.2% 195.7% 114.7%93.8% 88.0% 241.5% C0100M 173.6% 245.9% 195.6% 103.6% 110.6% 411.8%C0101M 138.2% 274.3% 174.8% 82.5% 123.4% 368.0% C0102M 131.8% 272.0%150.4% 78.6% 122.4% 316.6% C0104M 188.2% 238.9% 143.8% 99.5% 96.3%296.5% C0105M 198.1% 220.3% 73.1% 104.7% 88.8% 150.7% C0106M 171.8%240.7% 117.2% 102.5% 108.3% 246.7% C0108M 205.6% 258.5% 76.9% 108.7%104.1% 158.6% C0114M 114.0% 144.3% 35.9% 77.6% 71.4% 91.3% C0115M 177.2%226.8% 118.4% 105.7% 102.0% 249.3% C0116M 258.4% 302.8% 152.0% 136.6%122.0% 313.4% C0118M 166.2% 261.5% 79.2% 87.8% 105.4% 163.3% C0119M105.7% 167.8% 35.1% 71.9% 83.0% 89.3% C0124M 252.0% 305.1% 61.4% 133.2%122.9% 126.6% C0125M 168.6% 195.2% 159.7% 89.1% 78.6% 329.3% C0126M181.8% 265.3% 108.5% 108.5% 119.3% 228.4% C0128M 197.8% 286.0% 63.9%104.5% 115.2% 131.8% C0133M 139.4% 214.8% 72.4% 83.2% 96.6% 152.4%C0134M 158.5% 207.3% 46.6% 94.6% 93.3% 98.1% F-C0134 290.6% 378.9% 66.6%138.1% 111.4% 116.2% C0135M 161.3% 310.1% 113.3% 85.3% 124.9% 233.6%C0136M 176.8% 237.3% 74.5% 93.4% 95.6% 153.6% (P5) 0137M 180.8% 193.8%55.8% 95.6% 78.1% 115.1% (P7) C0142M 143.7% 192.5% 98.7% 97.8% 95.2%251.1% C0143M — — — — — — C0144M-2 — — — — — — C0145M-3 — — — — — —C0149M-2 — — — — — — C0150M-2 — — — — — — C0151M-2 — — — — — — C0152M-2— — — — — — C0153M-3 — — — — — — DAMGO 168.5% 266.1% 53.2% — — — Average

Example 2 FITC-NLX-Based FLNA Screening Assay

A. Streptavidin-Coated 96-Well Plates

Streptavidin-coated 96-well plates (Reacti-Bind™ NeutrAvidin™ Highbinding capacity coated 96-well plate, Pierce-ENDOGEN) are washed threetimes with 200 μl of 50 mM Tris HCl, pH 7.4 according to themanufacturer's recommendation.

B. N-biotinylated VAKGL pentapeptide (Bn-VAKGL) (SEQ ID NO: 1)

Bn-VAKGL peptide (0.5 mg/plate) is dissolved in 50 μl DMSO and thenadded to 4450 μl of 50 mM Tris HCl, pH 7.4, containing 100 mM NaCl andprotease inhibitors (binding medium) as well as 500 μl superblock in PBS(Pierce-ENDOGEN) [final concentration for DMSO: 1%].

C. Coupling of Bn-VAKGL Peptides to Streptavidin-Coated Plate

The washed streptavidin-coated plates are contacted with 5 μg/well ofBn-VAKGL (100 μl) for 1 hour (incubated) with constant shaking at 25° C.[50 μl of Bn-VAKGL peptide solution from B+50 μl binding medium, finalconcentration for DMSO: 0.5%]. At the end of the incubation, the plateis washed three times with 200 μl of ice-cold 50 mM Tris HCl, pH 7.4.

D. Binding of FITC-tagged naloxone [FITC-NLX] to VAKGL.

Bn-VAKGL coated streptavidin plates are incubated with 10 nM fluoresceinisothiocyanate-labeled naloxone (FITC-NLX; Invitrogen) in binding medium(50 mM Tris HCl, pH 7.4 containing 100 mM NaCl and protease inhibitors)for 30 minutes at 30° C. with constant shaking. The final assay volumeis 100 μl. At the end of incubation, the plate is washed twice with 100μl of ice-cold 50 mM Tris, pH 7.4. The signal, bound-FITC-NLX isdetected using a DTX-880 multi-mode plate reader (Beckman).

E. Screening of Medicinal Chemistry Analogs

The compounds are first individually dissolved in 25% DMSO containing 50mM Tris HCl, pH 7.4, to a final concentration of 1 mM (assisted bysonication when necessary) and then plated into 96-well compound plates.To screen new compounds, each compound solution (1 μl) is added to theBn-VAKGL coated streptavidin plate with 50 μl/well of binding mediumfollowed immediately with addition of 50 μl of FITC-NLX (total assayvolume/well is 100 μl). The final screening concentration for eachcompound is 10 μM.

Each screening plate includes vehicle control (total binding) as well asnaloxone (NLX) and/or naltrexone (NTX) as positive controls. Compoundsare tested in triplicate or quadruplicate. Percent inhibition ofFITC-NLX binding for each compound is calculated [(Total FITC-NLX boundin vehicle —FITC-NLX bound in compound)/Total FITC-NLX bound invehicle]×100%]. To assess the efficacies and potencies of the selectedcompounds, compounds that achieve approximately 60-70% inhibition at 10μl are screened further at 1 and 0.1 μM concentrations.

The results of this screening assay are shown in the table below.

FLNA Peptide Binding Assay Concentration of FLNA-binding FLNA-bindingCompound Compound 0.01 μM 0.1 μM 1 μM C0011 37.6% 41.4% 46.0% C002642.3% 44.8% 49.0% C0027 50.8% 61.2% 63.8% S-C0027 39.1% 46.5% 53.6%C0034-3 — — — C0037-2 — — — C0040 38.4% 46.3% 55.9% C0043 43.9% 51.3%58.0% C0044 37.3% 43.9% 50.6% C0045 39.1% 48.9% 53.7% C0046 30.8% 35.7%42.2% C0050 26.7% 34.5% 36.4% C0055 29.0% 34.9% 39.5% C0056 33.7% 38.9%41.4% C0060 60.3% 64.0% 68.0% C0086M 37.9% 48.1% 53.4% C0087M 51.6%57.9% 61.5% C0088M 40.1% 52.4% 56.1% C0089M 40.7% 46.1% 51.2% C0090M42.5% 52.5% 55.8% C0091M 38.1% 39.8% 46.3% C0093M 44.8% 49.9% 53.5%C0094M 43.0% 52.8% 57.5% C0095M 40.1% 46.6% 50.5% C0096M 43.0% 48.3%55.0% C0099M 46.9% 53.3% 56.0% C0100M 52.2% 58.2% 64.5% C0101M 50.5%56.4% 59.0% C0102M 52.3% 53.1% 56.6% C0104M 51.4% 54.1% 55.2% C0105M55.7% 62.0% 68.8% C0106M 45.8% 55.6% 58.9% C0108M 54.6% 61.4% 68.7%C0114M 57.1% 63.2% 66.7% C0115M 47.8% 57.8% 59.9% C0116M 53.9% 60.0%62.9% C0118M 56.6% 61.4% 62.4% C0119M 41.6% 55.5% 60.0% C0125M 54.2%59.7% 63.3% C0126M 50.7% 55.4% 67.3% C0128M 46.5% 54.4% 58.2% C0133M47.8% 54.9% 58.5% C0134M 55.7% 60.5% 61.9% F-C0134 37.4% 45.7% 53.1%C0135M 53.9% 55.1% 62.3% C0136M(P5) 46.7% 55.2% 58.2% C0137M(P7) 42.4%49.9% 61.2% C0142M 35.1% 39.4% 56.0% C0143M — — — C0144M-2 — — —C0145M-3 — — — C0149M-2 — — — C0150M-2 — — — C0151M-2 — — — C0152M-4 — —— C0153M-3 — — — Naloxone 40.61%  47.75%  51.54%  Average

Example 3 Tail-Flick Test

The mouse “tail flick” test was used to assay the relativeantinociceptive activity of compositions containing a compound to beassayed. This assay was substantially that disclosed by Xie et al., 2005J. Neurosci 25:409-416.

The mouse hot-water tail-flick test was performed by placing the distalthird of the tail in a water bath maintained at 52° C. The latency untiltail withdrawal from the bath was determined and compared among thetreatments. A 10 second cutoff was used to avoid tissue damage. Data areconverted to percentage of antinociception by the following formula:(response latency−baseline latency)/(cutoff−baseline latency)×100 togenerate dose-response curves. Linear regression analysis of the logdose-response curves was used to calculate the A₅₀ (dose that resultedin a 50% antinociceptive effect) doses and the 95% confidence intervals(CIs). Relative potency was determined as a ratio of the A₅₀ values. Thesignificance of the relative potency and the confidence intervals aredetermined by applying the t test at p<0.05.

To assess tolerance to the antinociceptive effect, the compound wasadministered twice daily for 7 days at an A₉₀ dose (dose that results ina 90% antinociceptive effect in the 52° C. warm-water tail-flick test),and the tail-flick test was performed daily after the a.m. dose. Asignificant reduction in tail-flick latency on subsequent days comparedto the Day 1 administration of the A₉₀ dose indicates antinociceptivetolerance. Assays were typically carried out separately using at least32 and 56 mg/kg dosages, with some assays also separately carried out at18 and 100 mg/kg.

Orally administered morphine exhibited an A₅₀ value of 61.8 (52.4-72.9)mg/kg, and a mean maximum antinociception amount of about 43% at 56mg/kg at about 20 minutes. Orally administered compound C0027 exhibiteda mean maximum antinociception amount of about 70% at 56 mg/kg at about20 minutes, whereas orally administered compound C0134M exhibited a meanmaximum antinociception amount of about 50% at 56 mg/kg at about 30minutes, compound C0066 exhibited a mean maximum antinociception amountof about 55% at 56 mg/kg at about 20 minutes, compound C0108M exhibiteda mean maximum antinociception amount of about 50% at 56 mg/kg at about20 minutes, compound C0090 exhibited a mean maximum antinociceptionamount of about 35% at 56 mg/kg at about 20 minutes, and compound C0089exhibited a mean maximum antinociception amount of about 30% at 56 mg/kgat about 20 minutes.

Example 4 Dependence test

On day 8, 16-20 hours after the last administration of an assaycomposition, animals were given naloxone to precipitate withdrawal (10mg/kg, s.c.) before being placed in an observation chamber for 1 hour. Ascale adapted from MacRae et al., 1997Psychobiology 25:77-82 was used toquantify four categories of withdrawal behaviors: “wet dog” shakes, pawtremors, mouth movements, and ear wipes. Scores are summed to yield atotal withdrawal score across the 1-hour test.

Example 5 Relative Gs/Go Switching

In this set of studies, the rat brain slice organotypic culture methodswere modified from those published previously (Adamchik et al., 2000Brain Res Protoc 5:153-158; Stoppini et al., 1991 J Neurosci Methods37:173-182). Striatal slices (200 μM thickness) were prepared using aMcllwain tissue chopper (Mickle Laboratory Engineering Co., Surrey, UK).Slices were carefully transferred to sterile, porous culture inserts(0.4 μm, Millicell-CM) using the rear end of a glass Pasteur pipette.Each culture insert unit contained 2 slices and was placed into one wellof the 12-well culture tray. Each well contain 1.5 ml of culture mediumcomposed of 50% MEM with Earl's salts, 2 mM L-glutamine, 25% Earl'sbalanced salt solution, 6.5 g/l D-glucose, 20% fetal bovine serum, 5%horse serum, 25 mM HEPES buffer, 50 mg/ml streptomycin and 50 mg/mlpenicillin. The pH value was adjusted to 7.2 with HEPES buffer.

Cultures were first incubated for 2 days to minimize the impact ofinjury from slice preparation. Incubator settings throughout theexperiment were 36° C. with 5% CO₂. To induce tolerance, culture mediumwas removed and the culture insert containing the slices was gentlyrinsed twice with warm (37° C.) phosphate-buffered saline (pH 7.2)before incubation in 0.1% fetal bovine serum-containing culture mediumwith 100 μM morphine for 1 hour twice daily (at 9-10 AM and 3-4 PM) for7 days.

Slices were returned to culture medium with normal serum after each drugexposure. Tissues were harvested 16 hours after the last drug exposureby centrifugation.

For determination of MOR-G protein coupling, slices were homogenated togenerate synaptic membranes. Synaptic membranes (400 μg) were incubatedwith either 10 μM oxycodone or Kreb's-Ringer solution for 10 minutesbefore solubilization in 250 μl of immunoprecipitation buffer (25 mMHEPES, pH 7.5; 200 mM NaCl, 1 mM EDTA, 50 μg/ml leupeptin, 10 μg/mlaprotinin, 2 μg/ml soybean trypsin inhibitor, 0.04 mM PMSF and mixtureof protein phosphatase inhibitors). Following centrifugation, striatalmembrane lysates were immunoprecipitated with immobilized anti-Gαs/olfor -Gαo conjugated with immobilized protein G-agarose beads. The levelof MOR in anti-Gasαolf or -Gαo immunoprecipitates was determined byWestern blotting using specific anti-MOR antibodies.

To measure the magnitude of MOR-mediated inhibition of cAMP production,brain slices were incubated with Kreb's-Ringer (basal), 1 μM DAMGO, 1 μMforskolin or 1 μM DAMGO+1 μM forskolin for 10 minutes at 37° C. in thepresence of 100 μM of the phosphodiesterase inhibitor IBMX. Tissues werehomogenized by sonication and protein precipitated with 1M TCA. Thesupernatant obtained after centrifugation was neutralized using 50 mMTris, pH 9.0. The level of cAMP in the brain lysate was measured by acAMP assay kit (PerkinElmer Life Science, Boston) according tomanufacturer's instructions.

Gs/Go-Coupled Condition Gs/olf Go Ratio Vehicle Average 330.7 1996.40.173 SEM 34.6 192.0 0.34 Oxycodone, 10 μM Average 1425.2 900.4 1.588SEM 77.8 26.2 0.103 C0011, 10 μM Average 534.3 1603.3 0.332 SEM 51.868.5 0.023 C0011, 100 μM Average 658.2 1598.8 0.420 SEM 34.2 114.9 0.030

Example 6 Carrageenan-Induced Acute Inflammatory Pain

To test the antinociceptive activity of the compounds under acuteinflammatory conditions, the latency to paw withdrawal from a noxiousthermal stimulus is determined before and 3 hours after injection of a50 μl solution of 2% carrageenan into the plantar surface of the hindpaw(Mogil et al. 1999 Pain 80:67-82). Animals are placed in plexiglas boxeson top of a glass plate maintained at 30° C. and allowed to habituatefor two sessions (−24 hours and −1 hour). Each habituation session lastsapproximately 45-60 minutes.

For baseline paw withdrawal latencies, an infrared heat source (UgoBasile model 37370) is applied from under the glass plate onto theplantar surface of the right hind paw with the focus of the light beamno larger than a 3- to 5-mm diameter. The time to withdrawal of the hindpaw from the heat source is recorded. A maximum cutoff of 30 seconds isused to prevent tissue damage. The intensity of the beam is set so thatbaseline latencies are approximately 15 seconds. The post-carrageenanbaseline is reestablished 3 hours after the carrageenan injections andonly animals with a significant decrease in the latency of hind pawwithdrawal from the thermal stimulus (thermal hypersensitivity) aretested. Animals are administered compounds, and hind paw withdrawallatencies are tested at various intervals after injection until the drugresponse falls below ˜20% MPE.

Antihyperalgesia (thermal hypersensitivity) and antinociception arecalculated as follows: percentage activity=100 [(test paw withdrawallatency−post-carrageenan baseline paw withdrawallatency)/(pre-carrageenan baseline paw withdrawallatency−post-carrageenan baseline paw withdrawal latency)].

Paw edema is determined by use of a plethysmometer (Ugo Basile) in themice undergoing the thermal latency testing. Paw volumes for the leftand right hind paw are measured at the conclusion of the thermal latencytesting (120 minutes after drug administration).

Example 7 LPS-Induced Cytokine Release from Primary Human Astrocytes

The following study was undertaken to investigate whether contemplatedcompounds and (+)NLX affect LPS-induced release of the pro-inflammatorycytokine (IL-1β, IL-6 and TNF-α) release from primary human astrocytes.

Concentrations used and duration of the treatments:

LPS: 1 mg/ml for 24 hours

-   (1) F-C0134: 100 fM, 10 pM, 1 nM, 100 nM: 2 hours prior to LPS and    continue for 24 hours-   (2) S-C0027: 100 fM, 10 pM, 1 nM, 100 nM: 2 hours prior to LPS and    continue for 24 hours

Experimental Design:

Primary astrocytes culture was prepared according to the provider'sinstructions (Lonza). The adherent astrocytes were trypsinized by 0.25%trypsin-EDTA, then collected and sub-cultured in 12-well plate (1.2ml/well). When the cells were 80-85% confluent, cells were treated in anincubator under 5% CO₂ for 2 hours with (1) 100 fM, 10 pM, 1 nM or 100nM F-C0134 and S-C0027, as well as (2) 10 pM and 1 nM (+)naloxone priorto addition of LPS (1 μg/ml). Vehicle groups were pretreated with 0.1%DMSO only. Following addition of LPS, incubation continued for 24 hours.Culture medium was used as the blank and the levels of cytokines, TNF-α,IL-6 and IL-1β in 200 μl of culture medium were determined. Each wellwas sampled twice.

To determine the effect of F-C0134, S-C0027 and (+)NLX on cytokinerelease, 0.5 μg/well of biotinated mouse monoclonal anti-TNF-α, -IL-6and -IL-1β were separately coated onto individual streptavidin-coatedplates (Reacti-Bind™ NeutrAvidin™ High binding capacity coated 96-wellplates), with different antibodies going into different wells. Plateswere washed 3 times with ice-cold 50 mM Tris HCl (pH 7.4) and incubatedat 30° C. with 200 μl medium derived from the above mentionedconditions. Plates were washed 3 times with ice-cold 50 mM Tris HCl (pH7.4) and incubated at 30° C. with 0.5 μg/well un-conjugated rabbitanti-anti-TNF-α, -IL-6 and -IL-1β for 1 hour. After two 1 minute washeswith 50 mM Tris HCl (pH 7.4), each well was incubated in 0.5 μg/wellFITC-conjugated anti-rabbit IgG (human and mouse absorbed) for 1 hour at30° C. Plates were washed twice with 200 μl ice-cold Tris HCl, pH 7.4and the residual FITC-Aβ₄₂ signals were determined by multimode platereader, DTX880 (Beckman).

TNF-α IL-6 IL-1β Compound Average % Average % Average % concentration(±SEM) (±SEM) (±SEM) F-C0134 100 fM 77.0 (2.8) 81.3 (2.4) 73.3 (5.3)  10pM 75.7 (1.8) 78.8 (1.5) 78.0 (5.5)  1 nM 77.4 (4.2) 78.2 (1.8) 77.7(2.9) 100 nM 75.4 (1.6) 76.3 (2.8) 73.2 (3.9) S-C0027 100 fM 77.7 (2.7)81.0 (1.7) 86.8 (2.0)  10 pM 73.6 (3.8) 75.6 (1.5) 85.8 (1.3)  1 nM 72.7(0.7) 77.4 (4.2) 81.9 (2.3) 100 nM 74.0 (1.7) 83.1 (2.1) 79.5 (2.4)(+)Naloxone  10 pM 58.6 (2.0) 56.9 (4.4) 75.9 (4.5)  1 nM  9.2 (6.0) 6.8 (4.3) 10.5 (4.8)

Compound Syntheses

A compound useful herein can be readily synthesized. An synthetic schemeis shown below that illustrates preparation of compounds containing asulfonyl linkage, a urea linkage, benzyl linkage and a carbonyl linkage.That scheme can be readily adapted for the preparation of compoundscontaining those same linkages in the configurations different fromthose shown, as well as for preparing different spiro ringconfigurations that utilize the same or alternative heteroatoms. Moredetailed syntheses are set out hereinafter.

Preparation of Compound 3-3

a. Preparation of compound 3-2

4-Methylbenzene-1-sulfonyl chloride (1.04 g, 5.49 mmol) was added to asolution of compound 3-1 (0.8 g, 5.23 mmol) in pyridine (20 mL) in anatmosphere of N₂ and the mixture was allowed to react overnight (about18 hours) at room temperature. Water was added and the resultingreaction mixture was extracted with CH₂Cl₂ 3 times. The combined organiclayers were washed with 3M HCl and brine and concentrated to givecompound 3-2 (0.78 g, yield: 59%, NMR confirmed).

b. Preparation of Compound 3-3

A solution of compound 3-2 (250 mg, 0.99 mmol), p-toluenesulfonic acidmonohydrate (20 mg) and 2-aminoethanol (5 mL) in ethanol (20 mL) wasstirred overnight (about 18 hours) at room temperature. The solvent wasremoved under reduced pressure and the residue was partitioned betweenethyl acetate and water. The organic layer was washed with water andbrine, dried with Na₂SO₄ and concentrated to give compound 3-3 (230 mg,yield: 80%, NMR confirmed) as a white solid.

Preparation of Compound 3-5

a. Preparation of Compound 3-4

To a solution of piperidin-4-one (0.47 g, 3.08 mmol) in pyridine (20 mL)was added 4-acetylaminobenzene sulfonyl chloride (0.6 g, 2.57 mmol). Themixture was stirred overnight (about 18 hours) at room temperature. Thenthe solvent was removed under reduced pressure. To the residue was addedCH₂Cl₂ (100 mL) and 2N HCl (50 mL). The organic layer was separated andwashed with 2N HCl (30 mL×2), then dried over Na₂SO₄ and concentrated togive the title compound as yellow solid. (0.4 g, yield: 52.6%).

b. Preparation of Compound 3-5

To a solution of compound 3-4 (0.55 g, 1.86 mmol) in ethanol (50 mL) wasadded p-toluenesulfonic acid monohydrate (50 mg) and 2-aminoethanol (0.5g, 8.2 mmol). The mixture was stirred overnight (about 18 hours) at 26°C. Then the solvent was removed under reduced pressure. To the residuewas added CH₂Cl₂ (100 mL) and saturated Na₂CO₃ (100 mL). The organiclayer was separated and washed with saturated Na₂CO₃ (50 mL×3), thendried over Na₂SO₄ and concentrated to give the crude product as whitepowder. (0.59 g, yield: 92.1%).

Preparation of Compound 3-7

a. Preparation of Compound 3-6

To a solution of compound I (150 mg, 1.11 mmol) in pyridine (4 mL) wastreated with 4-phenylnezenesulfonyl chloride (279.2 mg, 1.11 mmol). Themixture was stirred at room temperature overnight (about 18 hours). Tothe solution was added water and then extracted with dichloromethane (3times). The combined organic phase was washed with 3M HCl andconcentrated to give 205 mg of desired product as solid (¹H NMRconfirmed, 58.6% yield).

b. Preparation of Compound 3-7

To a solution of compound 3-6 (205 mg, 0.65 mmol) in ethanol (EtOH) (6mL) was treated with p-toluenesulfonic acid monohydrate (20 mg) andHOCH₂CH₂NH₂ (2 mL). The mixture was stirred at room temperatureovernight (about 18 hours). Then EtOH was removed under reducedpressure. The residue was partitioned between dichloromethane and water.The organic phase was washed by saturated aqueous NaHCO₃ and brine. Thenorganic layer was concentrated to give 202 mg of crude as white liquid(yield 87%).

Preparation of Compound 3-11

a. Preparation of Compound 3-10

4-Isopropylbenzene-1-sulfonyl chloride (0.13 mL, 0.7375 mmol) was addedto a solution of piperidin-4-one hydrochloride hydrate (100 mg, 0.7375mmol) in pyridine (3 mL) and the reaction mixture was stirred at roomtemperature for 3 hours. Water was added and the resulting reactionmixture was extracted with CH₂Cl₂ 3 times. The combined organic layerswere washed with 3M HCl and concentrated to give compound 3-10 (105 mg,yield: 50.7%, NMR confirmed) as a white solid.

b. Preparation of Compound 3-11

To a solution of compound 3-10 (200 mg, 0.71 mmol) in ethanol (EtOH) (6mL) was added p-toluenesulfonic acid monohydrate (15 mg) and2-aminoethanol (1.5 mL) and the reaction mixture was stirred overnight(about 18 hours) at room temperature. EtOH was removed under reducedpressure and the residue was partitioned between CH₂Cl₂ and water. Theorganic phase was washed with saturated aqueous NaHCO₃ and brine andconcentrated to give compound 3-11 (231 mg, yield: 100%) as a whitefoam.

Preparation of Compound 3-13

a. Preparation of Compound 3-12

A solution of compound 1 (300 mg, 2.21 mmol) in pyridine (8 mL) wasadmixed with 4-methoxy-sulfonylbenzene-1-sulfonyl chloride (0.34 mL,2.21 mmol). The mixture was stirred at room temperature for 3 hours. Tothe solution was added water and that composition was extracted withdichloromethane 3 times. The combined organic phase was washed with 3 MHCl and concentrated to give 335 mg of white solid (¹H NMR confirmed,56% yield).

a. Preparation of Compound 3-12

A solution of compound 3-12 (335 mg, 1.244 mmol) in ethanol (10 mL) wastreated with p-toluenesulfonic acid monohydrate (25 mg) and HOCH₂CH₂NH₂(2 mL). The mixture was stirred at room temperature overnight (about 18hours). The ethanol was removed under reduced pressure. The residue waspartitioned between dichloromethane and water. The organic phase waswashed by saturated NaHCO₃ and brine then concentrated to provide 380 mgof colorless oil (yield 97.7%).

b. Preparation of Compound 3-13

To a solution of compound 3-12 (335 mg, 1.244 mmol) in ethanol (10 mL)was added p-toluene-sulfonic acid monohydrate (25 mg) and 2-aminoethanol(2 mL) and the reaction mixture was stirred overnight (about 18 hours)at room temperature. Ethanol was removed under reduced pressure and theresidue was partitioned between CH₂Cl₂ and water. The organic phase waswashed with saturated NaHCO₃ and brine and concentrated to give compound3-13 (380 mg, yield: 97.7%) as a colorless oil.

Preparation of Compound 3-15

a. Preparation of Compound 3-14

To a solution of compound 3-1 (100 mg, 0.7375 mmol) in pyridine (3 mL)was added 4-trifluoromethoxy-benzene-1-sulfonyl chloride (192.38 mg,0.7375 mmol) and the reaction mixture was stirred at room temperaturefor 3 hours. Water was added and the resulting reaction mixture wasextracted with CH₂Cl₂ 3 times. The combined organic layers were washedwith 3M HCl and concentrated to give compound 3-14 (111 mg, yield:46.6%, ¹H-NMR confirmed) as a white solid.

b. Preparation of Compound 3-15

p-Toluenesulfonic acid monohydrate (10 mg) and 2-aminoethanol (1 mL)were added to a solution of compound 3-14 (111 mg, 0.343 mmol) inethanol (EtOH) (4 mL) and the reaction mixture was stirred at roomtemperature for 4 hours. EtOH was removed under reduced pressure and theresidue was partitioned between CH₂Cl₂ and water. The organic layer waswashed with saturated aqueous NaHCO₃ and brine and concentrated to givecompound 3-15 (128 mg of crude compound, NMR confirmed) as a lightyellow liquid.

Preparation of Compound 3-17

a. Preparation of Compound 3-16

2-Methyl-benzene-1-sulfonyl chloride (140.6 mg, 0.7375 mmol) was addedto a solution of compound 3-1 (100 mg, 0.7375 mmol) in pyridine (3 mL)and the reaction mixture was stirred overnight (about 18 hours) at roomtemperature. Water was added and the resulting reaction mixture wasextracted with CH₂Cl₂ 3 times. The combined organic layers were washedwith 3M HCl and concentrated to give compound 3-16 (104 mg, yield: 56%,¹H NMR confirmed) as a white solid.

b. Preparation of Compound 3-17

To a solution of compound 3-16 (104 mg, 0.41 mmol) in ethanol (EtOH) (4mL) was added p-toluenesulfonic acid monohydrate (10 mg) and2-aminoethanol (1 mL) and the reaction mixture was stirred overnight(about 18 hours) at room temperature. EtOH was removed under reducedpressure and the residue was partitioned between CH₂Cl₂ and water. Theorganic phase was washed with saturated aqueous NaHCO₃ and brine andconcentrated to give the crude compound 3-17 (120 mg, yield: 100%) as alight yellow liquid.

Preparation of Compound 3-25

a. Preparation of Compound 3-24

2-Cyanobenzenesulfonyl chloride (100 mg, 0.50 mmol) was added to asolution of piperidin-4-one (92 mg, 0.60 mmol) in pyridine (10 mL). Themixture was stirred at room temperature overnight (about 18 hours).Pyridine was removed by reduced pressure evaporation. The residue wasdissolved in CH₂Cl₂ (50 mL), and water (30 mL) was added. The CH₂Cl₂layer was separated and the water phase was extracted with CH₂Cl₂ (2×20mL). The organic layers were combined and washed with 3M HCl (20 mL×2).The organic layer was dried over anhydrous Na₂SO₄ and concentrated togive the title product as light-yellow oil (70 mg, yield: 53.4%,confirmed by MS).

b. Preparation of Compound 3-25

To a solution of compound 3-24 (35 mg, 0.13 mmol) in ethanol (10 mL) wasadded 2-aminoethanol (0.5 mL) and p-toluenesulfonic acid monohydrate (5mg). The mixture was stirred at 30° C. overnight (about 18 hours). Thesolvent was removed by evaporation under vacuum. To the residue wasadded CH₂Cl₂ (30 mL), then the CH₂Cl₂ layer was washed with saturatedNa₂CO₃ (15 mL×2) and water (20 mL×³), dried over Na₂SO₄ and concentratedto give the crude product as yellow oil (33 mg, yield: 80.5%, ¹H-NMRconfirmed).

Preparation of Compound 3-29

a. Preparation of Compound 3-28

3-Trifluoro-methoxybenzenesulfonyl chloride (287 mg, 1.1 mmol) was addedto a solution of compound 1 (150 mg, 1.1 mmol) in pyridine (7 mL). Themixture was stirred at room temperature overnight (about 18 hours).Water was added to the solution and then the solution was extracted withdichloromethane (3 times). The combined organic phase was washed with 3M HCl and concentrated to give 150 mg of the desired product as lightyellow solid (¹H NMR confirmed, 42% yield).

b. Preparation of Compound 3-29

A solution of compound 3-28 (140 mg, 0.46 mmol) in ethanol (EtOH) (6 mL)was treated with p-toluenesulfonic acid (15 mg) and HOCH₂CH₂NH₂ (1.5mL). The mixture was stirred at room temperature overnight (about 18hours). The EtOH was removed under reduced pressure. The residue waspartitioned between dichloromethane and water. The organic phase waswashed by saturated aqueous NaHCO₃ and brine. The organic layer wasconcentrated to give 145 mg of compound 3-29 as white liquid (¹H-NMRconfirmed, yield 85%).

Preparation of Compound 3-31

a. Preparation of Compound 3-30

Benzenesulfonyl chloride (200 mg, 1.13 mmol) was added to a solution ofpiperidin-4-one (208 mg, 1.36 mmol) in 20 mL of pyridine was addedbenzenesulfonyl chloride (200 mg, 1.13 mmol). The mixture was stirred atroom temperature overnight (about 18 hours). The pyridine was thenremoved by evaporation under vacuum. To the residue was added CH₂Cl₂ (50mL), then the CH₂Cl₂ layer was washed with 3M HCl (30 mL×3), dried overNa₂SO₄ and concentrated to give the crude product as a light yellowsolid (138 mg, yield: 51%).

b. Preparation of Compound 3-31

A solution of compound 3-30 (136 mg, 0.57 mmol), p-toluenesulfonic acidmonohydrate (15 mg) and 2-aminoethanol (2 mL) in ethanol (EtOH) (20 mL)was stirred overnight (about 18 hours) at room temperature. The solventwas removed by evaporation under vacuum. To the residue was added ethylacetate (50 mL) and water (50 mL). The ethyl acetate layer was washedwith water (30 mL×3). The water phase was washed with ethyl acetate (20mL). The combined organic layers were dried over Na₂SO₄, filtered andconcentrated to give the crude product (151 mg, yield: 92.5%). The crudeproduct was directly used in the next step.

Preparation of Compound 3-33

a. Preparation of Compound 3-32

3-Methyl-benzenesulfonyl chloride was added to a solution ofpiperidin-4-one hydrochloride monohydrate (159 mg, 1.03 mmol) in 10 mLpyridine (130 mg, 0.69 mmol). The mixture was stirred overnight (about18 hours) at room temperature. The pyridine was removed by evaporationunder vacuum. To the residue was added CH₂Cl₂ (50 mL), the CH₂Cl₂ layerwas washed with 3 M HCl (30 mL×3), dried over Na₂SO₄ and concentrated togive the crude product as light yellow solid (140 mg, yield: 80.5%).

b. Preparation of Compound 3-33

A solution of compound 3-32 (140 mg, 0.55 mmol), p-toluenesulfonic acid(15 mg) and 2-aminoethanol (2 mL) in ethanol (20 mL) was stirredovernight (about 18 hours) at room temperature. The solvent was removedby evaporation under vacuum. To the residue was added ethyl acetate (50mL) and water (50 mL). The ethyl acetate layer was washed with water (30mL×3), dried over Na₂SO₄ and concentrated to give the crude product as ayellow oil (170 mg).

Preparation of Compounds 3-35

a. Preparation of Compound 3-34

4-Methoxybenzoyl chloride (0.5 g, 2.93 mmol) was added to a solution ofpiperidin-4-one hydrochloride monohydrate (0.30 g, 1.95 mmol) inpyridine (20 mL). The reaction mixture was stirred at room temperatureovernight (about 18 hours). The reaction solvent was then removed underreduced pressure. The residue was dissolved in CH₂Cl₂ (50 mL), thenwashed with 3M HCl (50 mL×3). The organic layer was dried over Na₂SO₄and evaporated to give the title compound as a brown oil (330 mg, yield:61.5%, LC-MS confirmed).

b. Preparation of Compound 3-35

A solution of compound 3-34 (330 mg, 1.42 mmol), 2-aminoethanol (2 mL)and p-toluenesulfonic acid monohydrate (33 mg) in ethanol (20 mL) wasstirred at room temperature overnight (about 18 hours). The solvent wasthen removed by evaporation under reduced pressure. The residue wasdiluted with CH₂Cl₂ (50 mL), then washed with water (50 mL×3). Theorganic layer was dried over Na₂SO₄ and evaporated to give the crudeproduct as a yellow oil (360 mg, yield: 92.1%, ¹H-NMR and MS confirmed).

Preparation of Compound 3-37

a. Preparation of Compound 3-36

A solution of piperidine-4-one hydrochloride monohydrate (178 mg, 1.16mmol) in pyridine (20 ml) was treated with 4-methoxy-benzenesulfonylchloride (200 mg, 0.97 mmol). The mixture was stirred at roomtemperature overnight (about 18 hours). The pyridine was then removedunder reduced pressure. The residue was diluted with CH₂Cl₂ (50 mL),then washed with 3M HCl (30 mL×3). The organic layer was dried overanhydrous Na₂SO₄ and concentrated to give the product as a yellow solid(260 mg, yield: 100%, LC-MS confirmed).

b. Preparation of Compound 3-37

A solution of compound 3-36 (130 mg, 0.48 mmol), 2-aminoethanol (2 mL)and p-toluenesulfonic acid monohydrate (13 mg) in ethanol (20 mL) wasstirred at room temperature overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was dissolved in CH₂Cl₂ (50mL), then washed with saturated Na₂CO₃ (50 mL×2) and water (50 mL×2).The organic layer was then dried over Na₂SO₄ and concentrated to givethe product as a white colloid (118 mg, yield: 78.1%, LC-MS confirmed)

Preparation of Compound 3-38

p-Toluenesulfonic acid monohydrate (0.1 g) and 2-aminoethanol (2.45 g,40.2 mmol) were added to a solution of N-benzyl-4-piperidone (3.8 g,20.1 mmol) in ethanol (30 mL). The mixture was stirred at 30° C.overnight (about 18 hours). The solvent was removed under reducedpressure. To the residue was added CH₂Cl₂ (100 mL) and saturated Na₂CO₃(60 mL). The CH₂Cl₂ layer was separated and washed with saturated Na₂CO₃(50 mL×4). The organic layer was dried over Na₂SO₄ and concentrated togive the crude product as a brown oil (3 g, yield: 63.8%, ¹H NMRconfirmed).

Preparation of Compound 3-40

a. Preparation of Compound 3-39

To a solution of compound 3-38 (382 mg, 1.65 mmol) in pyridine (10 mL)was added p-acetyl-benzenesulfonyl chloride (300 mg, 1.37 mmol). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. To the residue was addedCH₂Cl₂ (50 mL), then the solution was washed with saturated Na₂CO₃aqueous (30 mL×3), dried over Na₂SO₄ and concentrated to give the crudeproduct as brown oil.

b. Preparation of Compound 3-40

To a solution of compound 3-39 (1.33 g, 3.2 mmol) in CH₂OH/CH₂Cl₂ (40/20mL) was added 10% Pd/C (270 mg). The mixture was stirred under H₂ atroom temperature for 24 hours. Thin-layer chromatography (TLC) indicatedthat no reaction had taken place. Then the Pd/C was replaced withPd(OH)₂/C, and the reaction was stirred under H₂ at room temperature andatmosphere pressure overnight (about 18 hours). TLC indicated that thereaction completed. The reaction mixture was filtrated and evaporated togive the crude product as light yellow solid (0.98 g, yield: 93.6%,LC-MS confirmed).

Preparation of Compound 3-44

a. Preparation of Compound 3-43

To a solution of piperidine-4-one hydrochloride monohydrate (0.52 g,3.38 mmol) in pyridine (10 mL), p-nitro-benzenesulfonyl chloride (0.5 g,2.26 mmol) was added. The reaction mixture was stirred overnight (about18 hours) at 30° C. and the solvent was removed under the reducedpressure. The residue was diluted with CH₂Cl₂ (30 mL), washed with 3NHCl (15 mL×3), the organic layer was dried, and evaporated to give thecrude compound as light yellow solid (200 mg, yield: 31%, LC-MSconfirmed).

b. Preparation of Compound 3-44

To a solution compound 3-43 (0.58 g, 2.04 mmol) in 20 mL of ethanol wasadded 2-aminoethanol (2 mL) and 4-methylbenzenesulfonic acid monohydrate(60 mg). The mixture was stirred at 25° C. overnight (about 18 hours).The solvent was removed under reduced pressure. The residue was dilutedwith CH₂Cl₂ (100 mL), washed with saturated Na₂CO₃ (100 mL×3) andsaturated NaHCO₃ (50 mL×3). The organic phase was dried over anhydrousNa₂SO₄ and concentrated to give the title compound as yellow solid (0.58g, yield: 90.6%)

Preparation of Compound C0008-2

a. Preparation of Compound C0008-1

To a solution of piperidin-4-one hydrochloride hydrate (343 mg, 2.23mmol) in pyridine (6 mL) was added 4-cyanobenzene-1-sulfonyl chloride(400 mg, 1.98 mmol). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. The residue was diluted with CH₂Cl₂ (50 mL) and washed with 1N HCl (50 mL×3). The organic phase was dried over anhydrous Na₂SO₄, andconcentrated to give the crude product as white solid (168 mg; yield:50%).

b. Preparation of Compound C0008-2

To the solution of compound C0008-1 (268 mg, 1.015 mmol) in ethanol (10mL) was added 2-aminoethanol (1.23 mL) and toluene-4-sulfonic acidmonohydrate (6 mg, 0.03 mmol). The mixture was stirred at roomtemperature overnight (about 18 hours). The solvent was removed underreduced pressure. The residue was diluted with CH₂Cl₂ (50 mL) and washedwith saturated aqueous NaHCO₃ (50 mL×6). The organic phase was driedover anhydrous Na₂SO₄ and concentrated to give compound C0008-2 asyellow oil (295 mg; yield: 94.7%).

Preparation of Compound C0009-2

a. Preparation of Compound C0009-1

4-(Methylsulfonyl)benzene-1-sulfonyl chloride (410 mg, 1.6 mmol) wasadded to a solution of piperidin-4-one hydrochloride hydrate (247 mg,1.6 mmol) in pyridine (10 mL). The mixture was stirred at roomtemperature overnight (about 18 hours). The solvent was removed underreduced pressure. To the residue was added CH₂Cl₂ (50 mL), then thesolution was washed with 1N HCl (20 mL×3), dried over Na₂SO₄ andconcentrated to give the title product 225 mg as yellow solid (yield:44%).

b. Preparation of Compound C0009-2

p-Toluenesulfonic acid monohydrate (4 mg) and 2-aminoethanol (0.8 mL,13.3 mmol) were added to a solution of compound C0009-1 (225 mg, 0.7mmol) in ethanol (6 mL). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. To the residue was added CH₂Cl₂ (70 mL) and washed withsaturated NaHCO₃ (25 mLx4), then dried over Na₂SO₄ and concentrated togive the product 205 mg as white solid (yield: 81%).

Preparation of Compound C0011-3

a. Preparation of Compound C0011-1

To a solution of N—BOC-piperidin-4-one (5 g, 23.2 mmol) andp-toluenesulfonic acid monohydrate (100 mg) in 50 mL of ethanol wasadded 2-aminoethanol (6 mL). The mixture was stirred at 25° C. overnight(about 18 hours). The solvent was removed under reduced pressure and theresidue was separated between 100 mL of dichloromethane and 100 mL ofsaturated Na₂CO₃, the organic layer was separated and washed with water(100 mL×3). Then the organic layer was dried over anhydrous Na₂SO₄ andconcentrated to give the crude product as yellow oil (5.9 g, yield:100%). ¹H NMR indicated that the crude product was a mixture containingthe title product, which was used directly in the next step withoutfurther purification.

b. Preparation of Compound C0011-2

4-Acetylbenzene-1-sulfonyl chloride (3 g, 13.7 mmol) was added to asolution of compound C0011-1 (5.3 g, 20.6 mmol) in 40 mL of pyridine.The mixture was stirred at room temperature overnight (about 18 hours).Then the solvent was removed under reduced pressure. The residue wasdiluted with CH₂Cl₂ (100 mL) and saturated Na₂CO₃ (100 mL), the organiclayer was separated and washed with saturated Na₂CO₃ (100 mL×3), andconcentrated to give the yellow oil, which was purified with silica gelcolumn to give the pure title compound (0.3 g, yield: 3.4%, ¹H NMR andLC-MS confirmed)

c. Preparation of Compound C0011-3

To a solution of compound C0011-2 (0.3 g, 0.71 mmol) in CH₂Cl₂ (10 mL)was added trifluoroacetic acid (2 mL). The mixture was stirred for 1hour at room temperature. CH₂Cl₂ (20 mL) was added, and saturated Na₂CO₃(30 mL) was added slowly with an ice-bath. The organic phase wasseparated, dried over anhydrous Na₂SO₄ and then concentrated to give thecrude product as yellow oil (160 mg, yield: 42%, LC-MS confirmed).

Preparation of Compound C0025-2

a. Preparation of Compound C0025-1

To a solution of piperidin-4-one hydrochloride monohydrate (1.8 g, 11.74mmol) in pyridine (30 mL) was added 4-bromobenzene-1-sulfonyl chloride(2 g, 7.83 mmol). The mixture was stirred overnight (about 18 hours) atroom temperature. The solvent was removed under reduced pressure. Theresidue was diluted with CH₂Cl₂ (100 mL), washed with 3N HCl (100 mL×2),dried over anhydrous Na₂SO₄ and concentrated to give the title compoundas a pale solid (1.3 g, yield: 52%, TLC confirmed).

b. Preparation of Compound C0025-2

A solution of C0025-1 (1.3 g, 4.09 mmol), 2-aminoethanol (5 mL) andp-toluenesulfonic acid monohydrate (130 mg) was stirred overnight (about18 hours) at 25° C. in 60 mL ethanol. The solvent was removed by reducedpressure evaporation. The residue was diluted with 200 mLdichloromethane, washed with water (100 mL×3) and saturated sodiumbicarbonate solutions (100 mL×3). Next, the organic layer was dried andconcentrated to get the product as a white solid. (1.44 g, yield: 97%,TLC confirmed).

Preparation of Compound C0026

Prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0027

a. Preparation of Compound 3-38

p-Toluenesulfonic acid monohydrate (100 mg) and 2-aminoethanol (5 mL)were added to a solution of N-benzyl-piperidin-4-one (10 g 52.8 mmol) in80 mL of ethanol. The mixture was stirred at 25° C. overnight (about 18hours). The solvent was removed under reduced pressure, the residue wasdiluted with 50 mL dichloromethane, and washed with saturated sodiumbicarbonate solutions (30 mL×3), saturated sodium carbonate (30 mL×3).The organic layer was dried and concentrated to get the product asyellow oil (11.5 g, yield: 93.8).

b. Preparation of Compound C0027-1

4-Methoxybenzene-1-sulfonyl chloride was added to the solution ofcompound 3-38 (1.37 g, 5.91 mmol) in pyridine (20 mL) (1.83 g, 8.85mmol). The reaction mixture was stirred overnight (about 18 hours) atroom temperature. The solvent was removed under reduced pressure. Theresidue (brown oil) was purified with silica gel column to give yellowfoam (410 mg, yield: 17%, confirmed by LC-MS).

c. Preparation of Compound C0027

To the solution of C0027-1 (410 mg, 1.02 mmol) in MeOH:CH₂Cl₂=2:1 (30mL), 10% Pd/C (0.2 g) was added and the reaction mixture was stirred atroom temperature overnight (about 18 hours) under H₂. The solvent wasfiltered to remove Pd/C. The solvent was removed under the reducedpressure to give the white foam as product (310 mg, yield: 98%,confirmed by LCMS).

¹H NMR (400 MHz, DMSO-d⁶) 6:7.79 (d, J=9.2 Hz, 2H), 7.15 (d, J=8.4 Hz,2H), 3.96 (t, J=6.4 Hz, 2H), 3.85 (s, 3H), 3.47 (t, J=6.4 Hz, 2H),3.28-3.24 (m, 2H), 2.86 (t, J=12 Hz, 2H), 2.46 (dt, J=13.6 Hz, 2H), 1.74(d, J=12.8 Hz, 2H); MS (ESI) calcd for C₁₄H₂₀N₂O₄S (m/z): 312.11. found:313.0 [M+1]⁺

Synthesis of S-C0027

a. Preparation of Compound B-1

2-Aminoethanethiol (25 g, 324.7 mmol) was added to a solution ofN-Boc-piperidin-4-one (32.4 g, 162.8 mmol) in ethanol (300 ml). Themixture was stirred at room temperature for 3 days. The solvent wasremoved under reduced pressure. The residue was diluted with CH₂Cl₂ (250mL) and washed with saturated aqueous Na₂CO₃ (150 mL×6). The organicphase was dried over anhydrous Na₂SO₄ and concentrated to obtain theproduct as a white solid (41.5 g, yield: 98.8%).

The structure was confirmed by MS (target M+1=259) and ¹H-NMR, shown asfollows:

¹H-NMR (400 MHz, CDCl₃): 3.79 (brs, 2H); 3.32 (t, J=6 Hz, 2H); 3.23-3.17(m, 2H); 2.97 (t, J=6.8 Hz, 2H); 1.83 (t, J=5.6 Hz, 4H); 1.44 (s, 9H).LC-MS (ESI) calcd for C₁₂H₂₂N₂O₂S (m/z): 258.38. found: 259.2 [M+1]⁺.

b. Preparation of Compound B-2

4-Methoxybenzene-1-sulfonyl chloride (26.8 g, 129.7 mmol) was added to asolution of B-1 (41.5 g, 160.85 mmol) in pyridine (300 ml). The mixturewas stirred at room temperature overnight (about 18 hours). Theresulting solution was evaporated and the residue was purified bychromatography to obtain compound B-2 as a white solid (54 g, yield:97%).

The structure was confirmed by LC-MS (target M+1-Boc (100)=329) and¹H-NMR, shown as follows:

¹H-NMR (400 MHz, CDCl₃): 7.80-7.76 (m, 2H); 6.99-6.95 (m, 2H); 4.24-4.14(m, 2H); 3.94-3.82 (m, 5H); 2.94-2.65 (m, 6H); 1.75 (brs, 2H); 1.46 (s,9H). LC-MS (ESI) calcd for C₁₉H₂₈N₂O₅S₂ (m/z): 428.57. found: 329.3[M−Boc+1]+.

c. Preparation of S-C0027

CF₃COOH (70 mL) was added to the solution of compound B-2 (28 g, 65.42mmol) in CH₂Cl₂ (700 mL). The mixture was stirred for 2 hours at roomtemperature. Then, the reaction mixture was adjusted to pH=10 withsaturated Na₂CO₃ followed by extraction with CH₂Cl₂:CH₃OH=10:1. Thecombined organic layer was dried over Na₂SO₄ and concentrated to obtainthe title product as white solid (18.6 g, yield: 86.67%).

The structure was confirmed by MS (target M+1=329), HPC and ¹H-NMR,shown as follows:

¹H-NMR (400 MHz, CDCl₃): 7.80-7.76 (m, 2H); 6.98-6.94 (m, 2H); 3.86 (s,3H); 3.84 (t, J=6 Hz, 2H); 3.15-3.11 (m, 2H); 2.86 (t, J=6.4 Hz, 2H);2.75-2.60 (m, 4H); 1.82-1.78 (m, 2H). LC-MS (ESI) calcd for C₁₄H₂₀N₂O₃S₂(m/z): 328.45. found: 329.3 [M+1]⁺.

Preparation of Compound C0029-2

a. Preparation of Compound C0029-1

4-Fluorobenzene-sulfonyl chloride (1 g, 5.14 mmol) was added to asolution of piperidine-4-one hydrochloride monohydrate (1.47 g, 9.57mmol) in pyridine (20 mL). The reaction mixture was stirred at roomtemperature overnight (about 18 hours). Next, the solvent was removedunder the reduced pressure and the residue was diluted with CH₂Cl₂ (20mL) and washed with 3 N HCl (15 mL×3). The organic layer was dried overanhydrous Na₂SO₄, filtered, and evaporated to give the crude compound aswhite solid (0.72 g, yield: 54.5%, ¹H-NMR confirmed).

b. Preparation of Compound C0029-2

A solution of compound C0029-1 (0.72 g, 2.8 mmol), 2-aminoethanol (0.26g, 4.2 mmol) and p-toluenesulfonic acid monohydrate (100 mg) in ethanol(20 mL) was stirred at 25° C. overnight (about 18 hours). The solventwas removed under the reduced pressure. The residue was diluted withCH₂Cl₂ (20 mL) and washed with NaHCO₃ solution (20 mL×3). The organiclayer was dried over anhydrous Na₂SO₄, filtered, and evaporated to givethe crude compound as white solid (0.81 g, yield: 96%, ¹H NMRconfirmed).

¹H-NMR (400 MHz, CDCl₃) δ: 7.81˜7.75 (m, 2H), 7.20˜7.14 (m, 2H), 3.67(t, J=6.4 Hz, 2H), 3.31˜3.26 (m, 2H), 3.12 (t, J=6.4 Hz, 2H), 2.97˜2.94(m, 2H), 1.76˜1.74 (m, 4H).

Preparation of Compound C0030

a. Preparation of Compound C0030-1

To a solution of piperidin-4-one hydrochloride monohydrate (594 mg, 3.9mmol) in 20 mL of pyridine was added 4-n-butylbenzenesulfonyl chloride(600 mg, 2.6 mmol). The mixture was stirred overnight (about 18 hours)at room temperature. The solvent was removed under reduced pressure. Theresidue was then diluted with 50 mL of dichloromethane, washed with 1Nhydrochloric acid (30 mL×3). Next, the organic layer was dried andconcentrated to give the crude product as a white solid (501 mg, yield:66%, ¹H NMR confirmed).

b. Preparation of Compound C0030-2

A solution of C0030-1 (500 mg, 1.7 mmol), 2-aminoethanol (5 mL) andp-toluenesulfonic acid monohydrate (100 mg) in 30 mL of ethanol wasstirred at 25° C. overnight (about 18 hours). The solvent was removed byreduced pressure evaporation. The residue was diluted with 50 mLdichloromethane, washed with water (50 mL×3) and saturated sodiumbicarbonate aqueous (50 mL×3). The organic layer was dried andconcentrated to give the product as a yellow solid (200 mg, yield: 89%,¹H NMR confirmed).

Preparation of Compound C0032-2

a. Preparation of Compound C0032-1

To a solution of piperidine-4-one hydrochloride monohydrate (3.15 g,20.51 mmol) in pyridine (30 mL), p-nitrobenzoyl chloride (2 g, 10.87mmol) was added. The reaction mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. The residue was diluted with CH₂Cl₂ (30 mL) and washed with 3NHCl (20 mL×3). The organic layer was dried over anhydrous Na₂SO₄,filtered, and evaporated to give the crude compound as a yellow solid(1.49 g, yield: 55.9%, confirmed by ¹H-NMR and LCMS).

b. Preparation of Compound C0032-2

A solution of compound C0032-1 (2 g, 8.06 mmol), 2-aminoethanol (0.73 g)and p-toluenesulfonic acid monohydrate (200 mg) in ethanol (40 mL) wasstirred at 25° C. overnight (about 18 hours). The solvent was removedunder reduced pressure. The residue was diluted with CH₂Cl₂ (30 mL) andwashed with NaHCO₃ (30 mL×3). Next, the organic layer was dried overanhydrous Na₂SO₄, filtered, and evaporated to give the crude compound asan orange solid. (2.2 g, yield: 93.7%, confirmed by ¹H-NMR and LCMS).

Preparation of Compound C0034-3

a. Preparation of Compound C0034-1

Cupric chloride (5 g) was added to a saturated solution of sulfurdioxide in CH₃COOH (200 mL) and sulfur dioxide gas (from the reaction ofNaHSO₄ and H₂SO₄). The gas was slowly bubbled into the solution for 4hours until the solution became blue-green. Next,4-amino-benzene-1-sulfonamide (20 g, 116 mmol) was added to a solutionof concentrated HCl (40 mL) and H₂O (50 mL) with stirring for 1 hour at0° C. To this mixture was added a solution of sodium nitrate (8 g, 116mmol) at such a rate of addition that the temperature did not rise above0° C. The mixture was stirred for 0.5 hours then quenched with theSO₂/CuCl₂ solution made earlier. The mixture was then stirred for 1 hourat room temperature. Next, H₂O (500 mL) was added, and stirringcontinued for an additional 30 minutes. The product was collected bysuction filtration, washed with H₂O, dried in vacuo at 60° C. to givethe title product as a light yellow solid (LC-MS confirmed). Afterdrying, about 10 g crude product as a light yellow solid was obtained(10 g, yield: 33%, confirmed by LC-MS).

b. Preparation of Compound C0034-2

Compound C0034-1 (2.00 g, 7.8 mmol) was added to a solution ofpiperidine-4-one hydrochloride monohydrate (1.4 g, 9.4 mmol) in 30 mLpyridine. The mixture was stirred overnight (about 18 hours) at roomtemperature. The solvent was removed under reduced pressure and theresidue was diluted with CH₂Cl₂. The crude product was washed with 2NHCl (50 mL×3). The aqueous layer was extracted with CH₂Cl₂. The organicphase was combined and concentrated to give the crude product as a lightyellow solid (0.65 g, yield: 37%, TLC confirmed).

c. Preparation of C0034-3

To a solution of compound C0034-2 (0.5 g, 1.58 mmol) in 10 mL ethanolwas added ethanolamine (5 mL) and 4-methylbenzenesulfonic acidmonohydrate (0.1 g). The mixture was stirred overnight (about 18 hours)at 25° C. Then the solvent was removed under reduced pressure. Theresidue was diluted with CH₂Cl₂ (100 mL), and washed with saturatedNaHCO₃ (50 mL×6), there was much dissolved solid. Then the organic phasewas dried over anhydrous Na₂SO₄ and concentrated to give few yellowsolid. The aqueous layer was filtered to provide a white solid. Thewhite solid was confirmed to be the product, which was purified withsilica gel column to give the pure product as white solid (0.25 g,yield: 43.9%, ¹H NMR confirmed).

Preparation of Compound C0037-2

a. Preparation of Compound C0040-1

To the solution of piperidin-4-one hydrochloride monohydrate (200 mg,1.3 mmol) in pyridine (20 mL) was added 4-nitrobenzene-1-sulfonylchloride (262 mg, 1.18 mmol). The mixture was stirred overnight (about18 hours) at room temperature. The solvent was removed under reducedpressure. The residue was diluted with 50 mL dichloromethane and washedwith 0.5 M HCl (50 mL×3). The organic layer was dried and evaporated togive the product as white solid. (180 mg, yield: 41.2%, confirmed by¹H-NMR.

b. Preparation of Compound C0037-2

3-Aminopropanol (0.5 mL, 6.54 mmol) and p-toluenesulfonic acidmonohydrate (30 mg) were added to the solution of compound C0040-1 (200mg, 0.7 mmol) in 8 mL ethanol. The mixture was stirred at 25° C.overnight (about 18 hours). Then the solvent was removed by evaporationunder reduced pressure. The residue was diluted with CH₂Cl₂ (20 mL),washed with saturated Na₂CO₃ (20 mL×3) and saturated NaHCO₃ (20 mL×3).Then the organic phase was dried over anhydrous Na₂SO₄ and concentratedto give the title compound as yellow solid (0.21 g; yield: 90.4%; LC-MS&¹H NMR confirmed, HPLC 96.7%).

Preparation of Compound C0040

Prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0044

a. Preparation of Compound C0040-1

To the solution of piperidin-4-one hydrochloride monohydrate (200 mg,1.3 mmol) in pyridine (20 mL) was added 4-nitrobenzene-1-sulfonylchloride (262 mg, 1.18 mmol). The mixture was stirred overnight (about18 hours) at room temperature. The solvent was removed under reducedpressure. The residue was diluted with 50 mL dichloromethane and washedwith 0.5 M HCl (50 mL×3). The organic layer was dried and evaporated togive the product as white solid. (180 mg, yield: 41.2%, confirmed by¹H-NMR).

b. Preparation of Compound C0044

To the solution of C0040-1 (170 mg, 0.5 mmol) and p-toluenesulfonic acidmonohydrate (1.7 mg) in ethanol (10 mL) was added cysteamine (46 mg, 0.6mmol). The mixture was stirred overnight (about 18 hours) at 25° C. Thesolvent was removed under reduced pressure. The residue was diluted withdichloromethane (50 mL) and washed with saturated Na₂CO₃ solution (40mL×3). The organic layer was dried and evaporated to give the crudeproduct as yellow solid. The solid was washed with 1M HCl. The mixturewas filtered to get the filtration as white solid. The solid was washedwith saturated Na₂CO₃ solution and extracted with dichloromethane. Theorganic layer was dried and evaporated to give the product as whitesolid. (120 mg, yield: 57.9%, confirmed by LCMS).

Preparation of Compound C0046

a. Preparation of Compound C0046-1

4-Acetylbenzene-1-sulfonyl chloride (2 g, 9.1 mmol) was added to asolution of C0011-1 (2.63 g, 10.9 mmol) in pyridine (20 mL). The mixturewas stirred at room temperature overnight (about 18 hours). The solventwas removed under reduced pressure. The residue was diluted with CH₂Cl₂(100 mL) and washed with saturated NaHCO₃ (100 mL×3). The organic phasewas dried over anhydrous Na₂SO₄ and concentrated to give the crudeproduct as yellow solid, which was purified with silica gel column(eluted with CH₂Cl₂:CH₃OH-80:1) to give the title compound as a whitesolid (3.3 g, yield: 71.7%, ¹H NMR confirmed).

b. Preparation of Compound C0046

To the solution of C0046-1 (3.26 g, 11.48 mmol) in dichloromethane (DCM)(20 mL) was add CF₃COOH (5 mL). The mixture was stirred overnight (about18 hours) at room temperature. Thin-layer chromatography indicated thatthe material reacted completely. DCM (30 mL) was added and the organiclayer was washed with saturated sodium carbonate solution (50 mL×3).Then the organic layer was dried and evaporated to get yellow oil, whichwas purified with silica gel column to give the product as yellow oil(1.1 g, yield: 44%, confirmed by LCMS, MS and ¹H NMR, HPLC: 98.2%).

Preparation of Compound C0049-2

a. Preparation of Compound C0049-1

3,4-Dimethoxy-benzene-1-sulfonyl chloride (500 mg, 2.1 mmol) was addedto a solution of piperidin-4-one hydrochloride monohydrate (486 mg, 3.2mmol) in 20 mL pyridine. The mixture was stirred overnight (about 18hours) at room temperature. Then the solvent was removed under reducedpressure. The residue was diluted with dichloromethane (50 mL) andwashed with 1 N HCl (30 mL×3). The organic layer was dried andconcentrated to give the product as yellow solid (260 mg, yield: 41.2%,confirmed by LCMS).

b. Preparation of Compound C0049-2

p-Toluenesulfonic acid monohydrate (26 mg) and 2-aminoethanol (5 mL)were added to a solution of compound C0049-1 (260 mg, 0.87 mmol) in 10mL of ethanol and stirred at 25° C. overnight (about 18 hours). Thesolvent was removed by reduced pressure evaporation. The residue wasdiluted with 50 mL of dichloromethane, and then washed with saturatedsodium bicarbonate solutions (20 mL×3). The organic layer was dried andconcentrated to give the product as yellow solid (297 mg, yield: 100%,confirmed by ¹H NMR).

Preparation of Compound C0050

a. Preparation of Compound C0040-1

To a solution of piperidin-4-one (200 mg, 1.2 mmol) in pyridine (20 mL)was added 4-nitrobenzene-1-sulfonyl chloride (262 mg, 1.18 mmol). Themixture was stirred overnight (about 18 hours) at room temperature. Thesolvent was removed under reduced pressure. The residue was diluted with50 mL dichloromethane and washed with 0.5 M HCl (50 mL×3). The organiclayer was dried and evaporated to give the product as white solid. (180mg, yield: 41.2%, confirmed by ¹H NMR).

b. Preparation of Compound C0044

To a solution of 4-nitrobenzene-1-sulfonyl chloride (170 mg, 0.6 mmol)in 20 mL ethanol was added 2-mercaptoethylamine (70 mg, 0.9 mmol) andp-toluenesulfonic acid (1.7 mg). The mixture was stirred overnight(about 18 hours) at 25° C. The solvent was removed and the residue wasdiluted with 50 mL dichloromethane, washed with saturated Na₂CO₃solutions (30 mL×3). The organic layer was separated, dried, andconcentrated to give the crude product as yellow solid, which waspurified with silica gel column to give the title product as white solid(100 mg, yield: 48.7%)

c. Preparation of Compound C0050

To the solution of C0044 (120 mg, 0.35 mmol) and triethylamine (106 mg,1.05 mmol) in dry dichloromethane (DCM) (20 mL) was added dropwiseacetyl chloride (55 mg, 0.7 mmol). The mixture was stirred overnight(about 18 hours) at room temperature. The mixture was washed with water,purified by silica gel column chromatography and then purified bypreparative thin-layer chromatography to give a white solid, but HPLCshowed it was not pure enough. The partially purified product wasfurther purified by silica gel column chromatography (ethylacetate:petroleum ether=3:1) to give the pure product as white solid (13mg, yield: 9.7%, confirmed by LCMS, ¹H-NMR and MS, HPLC:97.3%).

Preparation of Compound C0053-3

a. Preparation of Compound C0053-1

To a solution of 4-acetylbenzoic acid (250 mg, 1.52 mmol) in drydichloromethane (20 mL) and dimethylformamide (0.1 mL) was addeddropwise oxalyl chloride (570 mg, 4.5 mmol) at 0° C. After addition, themixture was stirred for 2 hours at room temperature. The solvent andexcess oxalyl chloride was removed by reduced pressure evaporation togive the product as a yellow solid (270 mg, yield: 97%, confirmed byLCMS dissolved with methanol.

b. Preparation of Compound C0053-2

To a solution of C0011-1 (727 mg, 23 mmol) and diisopropyl ethyl amine(1 mL) in dry dichloromethane (DCM) (20 mL) was added C0053-1 (500 mg,2.74 mmol solution in 20 mL dry DCM) dropwise at 0° C. The mixture wasstirred at room temperature for 3 days. The mixture was then washedthree times with water (50 mL), the organic layer was dried thenevaporated to get the product as brown oil (1.28 g, yield: 100%,confirmed by LCMS).

c. Preparation Compound C0053-3

A solution of C0053-2 (1 g, 2.58 mmol) and CF₃COOH (5 mL) indichloromethane (20 mL) was stirred overnight (about 18 hours) at roomtemperature. The mixture was washed with a saturated Na₂CO₃ solution,the organic layer was dried and evaporated to give the crude product asa brown oil. The crude product was purified on a silica gel columnchromatography to provide the purified product as a brown oil (360 mg,yield: 48.3%, confirmed by LCMS.)

Preparation of Compound C0055

a. Preparation of Compound C0055-1

To compound C0011-1 (1 g, 4.13 mmol) in dichloromethane (15 mL) wasadded triethylamine (1.72 mL, 12.39 mmol) and CBz-Cl (0.88 mL, 6.19mmol) in a salt-ice bath. The reaction mixture was stirred overnight(about 18 hours) at room temperature. The solvent was removed underreduced pressure. The residue was diluted with dichloromethane (30 mL),and washed with water (20 mL×2). The organic layer was dried andevaporated to give the crude product as light yellow oil (1.4 g, yield:91%, confirmed by ¹H NMR.)

b. Preparation of Compound C0055-2

A solution of C0055-1 (1.4 g, 3.72 mmol) and trifluoroacetic acid (3 mL)in dichloromethane (15 mL) was stirred overnight (about 18 hours) atroom temperature. The mixture was quenched with saturated Na₂CO₃solution, diluted with CH₂Cl₂ (20 mL) and washed with saturated Na₂CO₃solution (15 mL×2). The aqueous layer was extracted with CH₂Cl₂ (20mL×3). The organic layer was combined, dried over anhydrous Na₂SO₄ andevaporated to give the crude product as orange oil (0.88 g, yield: 86%,confirmed by LCMS and ¹H NMR).

c. Preparation of Compound C0055-3

To the solution of C0055-2 (0.34 g, 1.25 mmol) and diisopropylethylamine(0.46 mL, 2.81 mmol) in dichloromethane (10 mL), was added4-acetylbenzene-1-sulfonyl chloride (0.41 g, 1.87 mmol). The mixture wasstirred overnight (about 18 hours) at room temperature. The solvent wasremoved under reduced pressure. The mixture was diluted with CH₂Cl₂ (20mL) and washed with NaHCO₃ (15 mL×2) and water (15 mL). The organiclayer was dried and evaporated to give the crude compound as yellowsolid, which was purified with silica gel column to give the pureproduct as white solid (290 mg, yield: 52%, ¹H NMR and LCMS confirmed).

d. Preparation of Compound C0055-4

To the solution of C0055-3 (100 mg, 0.22 mmol) in methanol (10 mL), wasadded Pd(OH)₂/C (20 mg). The mixture was stirred overnight (about 18hours) at room temperature under H₂. Thin-layer chromatography showedthat the reaction was not completed, so a little more Pd(OH)₂/C wasadded, then stirred overnight (about 18 hours) under H₂. The mixture wasfiltered; the solution was evaporated to give the crude product as whitesolid (65 mg, yield: 91%, confirmed by LCMS, ¹H NMR.

Preparation of C0056

a. Preparation of C0056-1

To a solution of C0011-1 (500 mg, 2.06 mmol) in dry CH₂Cl₂ (10 mL) wasadded diisopropyl ethyl amine (0.68 mL, 4.12 mmol) and methane sulfonylchloride (0.24 mL, 3.1 mmol). The reaction mixture was stirred overnight(about 18 hours) at room temperature. The mixture was washed withsaturated NaHCO₃ (2×20 mL) and water (1×20 mL). The organic layer wasdried and concentrated to give the crude compound as orange oil (yield:650 mg, 99%, confirmed by ¹H NMR).

b. Preparation of C0056-2

To the solution of C0056-1 (650 mg, 2.04 mmol) in CH₂Cl₂ (8 mL),trifluoroacetic acid (2 mL) was added. The reaction mixture was stirredat room temperature overnight (about 18 hours). The mixture was quenchedwith sat Na₂CO₃ solution, diluted with CH₂Cl₂ (20 mL) and washed withNa₂CO₃ solution (15 mL×2). The aqueous layer was extracted with CH₂Cl₂(20 mL×4). The organic layer was dried over anhydrous Na₂SO₄ andevaporated to give the crude product as brown solid (178 mg, yield: 40%,confirmed by LC/MS).

c. Preparation of C0056

4-Acetylbenzene-sulfonyl chloride (0.26 g, 1.21 mmol) was added to thesolution of C0056-2 (178 mg, 0.81 mmol) in pyridine (10 mL). The mixturewas stirred overnight (about 18 hours) at room temperature. The solventwas removed under reduced pressure. The residue was diluted with CH₂Cl₂(30 mL) and washed with 1N HCl (20 mL×2) and water (20 mL). The organiclayer was dried and evaporated to give the crude compound as yellowsolid. Which was purified with silica gel column to give the pureproduct as white foam (60 mg, yield: 18.5%, ¹H NMR and MS confirmed,HPLC 98%).

¹H-NMR (400 MHz, CDCl₃) δ:8.09 (d, J=8.4 Hz, 2H), 7.86 (d, J=8.4 Hz,2H), 3.93 (t, J=6 Hz, 2H), 3.81 (dd, J=8.8 Hz, 1.6 Hz, 2H), 3.53 (t, J=6Hz, 2H), 2.94 (s, 3H), 2.81 (s, 3H), 2.66-2.49 (m, 4H), 1.74 (d, J=11.6Hz, 2H). MS (ESI) calcd for C₁₆H₂₂N₂O₆S₂ (m/z): 402.09. found:403.3[M+1]⁺, 425.2[M+23]⁺.

Preparation of Compound C0058

a) Preparation of Compound C0058-1

To the C0011-1 (600 mg, 2.48 mmol) and diisopropylethylamine (0.82 mL,4.96 mmol) in dichloromethane (DCM) (10 mL), ethylsulfonyl chloride (478mg, 3.72 mmol) was added. The reaction mixture was stirred overnight(about 18 hours) at room temperature. The reaction mixture was dilutedwith DCM (50 mL) and washed with saturated with aqueous Na₂CO₃ (30mL×3). The organic layer was dried over anhydrous Na₂SO₄ andconcentrated to give the crude product as brown-black oil. LC-MSindicated that there was no the desired product peak. Purification withsilica gel could not give the pure product.

Run#2: Pyridine ethyl sulfonyl chloride (0.3 mL, 3.17 mmol) was addeddropwise to the solution of N-Boc-piperidin-4-one (500 mg, 2.06 mmol) in15 mL of pyridine. The mixture was stirred at room temperature overnight(about 18 hours).

Preparation of Compound C0059

a. Preparation of C0059-1

Run#1: Isopropylsulfonyl chloride (0.35 mL, 3.09 mmol) was added To theC0011-1 (500 mg, 2.06 mmol) and diisopropylethylamine (1.02 mL, 6.18mmol) in dichloromethane (10 mL). The reaction mixture was stirredovernight at room temperature. The mixture was diluted withdichloromethane and washed with NaHCO₃ (20 mL×2). The organic layer wasdried and evaporated to give the crude product as dark yellow oil. Afterchecking LC-Ms, the peak of the product could not be observed, and TLCwas showed it was complicated

Run#2: Isopropyl sulfonyl chloride (0.28 mL, 2.47 mmol) was addeddropwise to the solution of C0011-1 (500 mg, 2.06 mmol) in pyridine (20mL). The mixture was stirred at room temperature overnight. The solventwas removed by reduced pressure evaporation. The residue was dilutedwith 50 mL of dichloromethane and washed with 0.5N HCl (40 mL×3). Theorganic layer was dried and evaporated to give the crude product asyellow solid. LCMS showed there was no the desired peak.

Preparation of Compound C0060

a. Preparation of Compound C0060-1

2-Mercaptoethylamine (100 mg, 1.3 mmol) was added to the solution ofpiperidin-4-one hydrochloride monohydrate (200 mg, 1.3 mmol) in ethanol(10 mL). The mixture was stirred overnight (about 18 hours) at 25° C.The solvent was removed under reduced pressure to give the product asyellow foam (310 mg, LC-MS showed there was the desired product peak.The crude was used directly in the next step.

b. Preparation of Compound C0060

4-Acetylbenzenesulfonyl chloride (0.348 g, 1.59 mmol) was added to thesolution of C0060-1 (0.31 g, 1.59 mmol) in 10 mL of pyridine. Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed by reduced pressure evaporation, the residue wasdiluted with 50 mL dichloromethane, and then washed with 1M HCl (30mL×3), and then the organic layer was dried with anhydrous sodiumsulfate and concentrated to give the crude product as yellow oil (0.1g). The crude product was further purified with silica gel column givingthe title compound C0060 as white solid (200 mg, yield: 37%, HPLC:95.99%, ¹H NMR and MS confirmed).

¹H NMR (400 MHz, CDCl₃) δ: 8.11 (d, J=8.4 Hz, 2H), 7.88 (d, J=8.4 Hz,2H), 3.46˜3.41 (m, 2H), 3.28 (t, J=6 Hz, 2H), 3.01˜2.93 (m, 4H), 2.68(s, 3H), 2.04˜1.99 (m, 4H), 1.67 (brs, 1H). MS (ESI) calcd forC₁₅H₂₀N₂O₃S₂ (m/z): 340.09. found: 341.1[M+1]⁺.

Preparation of Compound C0062-3

a. Preparation of Compound C0062-1

CuCl₂ (2.5 g) was added to the saturated solution of SO₂ (get from thereaction of NaHSO₃ and H₂SO₄) in glacial acetic acid (200 mL) and SO₂gas was slowly bubbled into the solution for 2 hours.3,4,5-Trimethoxyaniline (10 g, 54.6 mmol) was added to the solution ofconcentrated HCl (40 mL) and H₂O (50 mL) and the mixture was stirred for1 hour at 0° C. To this solution was added a solution of NaNO₂ (3.77 g,54.6 mmol) in H₂O (20 mL) at such a rate that the temperature did notrise above 0° C. The mixture was stirred for 0.5 hours and then addeddropwise to the SO₂ and CuCl₂ saturated solution before. The reactionwas then stirred for 1 hour. H₂O (1000 mL) was added and continuedstirring for 0.5 hours. Then the product was collected by suctionfiltration, washed with H₂O, and dried in vacuum at 50° C.

b. Preparation of Compound C0062-2

To a solution of piperidin-4-one hydrochloride (229 mg, 1.68 mmol) inpyridine (10 mL) was added compound C0062-1 (300 mg, 1.12 mmol). Thereaction mixture was stirred overnight (about 18 hours) at roomtemperature. To the reaction mixture 20 mL water was added, and thereaction mixture was extracted with dichloromethane three times. Theorganic layers was combined and was washed with 1N HCl two times andbrine one times, dried, concentrated under vacuum to afford 279 mg ofcrude product (yield: 75.7)

c. Preparation of Compound C0062-3

To a solution of C0062-2 (279 mg, 0.848 mmol) in ethanol (5 mL) wasadded compound p-toluenesulfonic acid monohydrate (4.51 mg, 0.024 mmol)and 2-aminoethanol (0.9 mL). The reaction mixture was stirred overnight(about 18 hours) at room temperature. The reaction mixture was dissolvedin CH₂Cl₂ (30 mL) and washed with aqueous of NaHCO₃ and brine, dried,concentrated under vacuum to afford 226 mg of crude product. (yield:71.6%).

Preparation of Compound C0065-9

a. Preparation of Mixture C0065-1 and C0065-6

A suspension of phthalimide (7.35 g, 50 mmol) in epichlorohydrin (15.7mL, 200 mmol) was boiled under reflux for 10 hours. The mixture wasallowed to cool and was concentrated producing a brown oil. The crudeproduct was purified by silica gel chromatography to obtain 4.7 g ofC0065-6 as white solid (yield: 46%) and 4.1 g of C0065-1 as white solid.

b. Preparation of Compound C0065-2

The mixture of compound C0065-1 and C0065-6 (3.1 g, 15 mmol) and aqueousdimethylamine (10.3 g) was stirred overnight (about 18 hours) at roomtemperature. Thin-layer chromatography suggested the reaction complete.The reaction mixture was washed with CH₂Cl₂ for 3 times, and the waterwas removed under reduced pressure to obtain the crude product ofC0065-2.

c. Preparation of Compound C0065-3

The compound C0065-2 was added into 20% hydrochloric acid (20 mL) andrefluxed for 4 hours. Then the mixture was cooled to room temperature,phthalic acid was separated. The aqueous solution was washed by etherand concentrated. The residue was dissolved in NaOH (20%) and extractedwith CH₂Cl₂ for 3 times. The combined organic layers were dried overNa₂SO₄ and concentrated to obtain 1.1 g of the product as yellow oil(yield: 62%).

d. Preparation of Compound C0065-7

To a solution of N-Boc-piperidin-4-one (420 mg, 2.1 mmol) in ethanol (4mL) was added compound C0065-3 (500 mg, 4.2 mmol). The mixture wasstirred overnight (about 18 hours) at room temperature. The solvent wasremoved under reduced pressure. The residue was diluted with CH₂Cl₂ (30mL) and washed with saturated aqueous Na₂CO₃ (30 mL×6). The organicphase was dried over anhydrous Na₂SO₄, then concentrated to give 600 mgof compound C0065-4 as a yellow oil (yield: 95%)

e. Preparation of Compound C0065-8

4-Acetylbenzene-1-sulfonyl chloride (200 mg, 0.9 mmol) was added to asolution of compound C0065-7 (600 mg, 2.0 mmol) in pyridine (4 mL). Themixture was stirred at room temperature for 4 hours. The solvent wasremoved under reduced pressure. The residue was purified with silica gelto obtain 220 mg of the title compound as a white solid (yield: 50%)

f. Preparation of Compound C0065-9

To the solution of compound C0065-8 (220 mg, 0.45 mmol) in 3 mL CH₂Cl₂was add 0.5 mL CF₃COOH and the mixture was kept stirring for 1 hour atroom temperature. To the mixture was added 30 mL of CH₂Cl₂ and washedwith saturated sodium carbonate solution (30 mL×3). The organic layerwas dried and concentrated to get the crude product 150 mg as whitesolid.

Preparation of Compound C0068-2

a. Preparation of Compound C0068-1

Piperidin-4-one hydrochloride hydrate (92 mg, 0.6 mmol) in pyridine (2mL) was treated with 4-bromo-2-(trifluoromethyl)benzene-1-sulfonylchloride (194 mg, 0.6 mmol). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. To the residue was added CH₂Cl₂ (50 mL), the solution waswashed with 1N HCl (20 mL×3), dried over Na₂SO₄, and concentrated togive the title product as yellow solid (117 mg; yield: 50.5%).

b. Preparation of Compound C0068-2

p-Toluenesulfonic acid monohydrate (1.59 mg) and 2-aminoethanol (0.32mL, 5.54 mmol) were added to a solution of compound C0068-1 (107 mg,0.28 mmol) in ethanol (4 mL). The mixture was stirred at roomtemperature overnight (about 18 hours). Then the solvent was removedunder reduced pressure. To the residue was added CH₂Cl₂ (70 mL) andwashed with saturated NaHCO₃ (25 mL×4), then dried over Na₂SO₄ andconcentrated to give the product as yellow oil (110 mg, yield: 91.6%).

Preparation of Compound C0071-2

a. Preparation of Compound C0071-1

Piperidin-4-one hydrochloride hydrate (95 mg, 0.62 mmol) in pyridine (3mL) was treated with 2-bromo-4-(trifluoromethyl)benzene-1-sulfonylchloride (200 mg, 0.62 mmol). The mixture was stirred at roomtemperature overnight (about 18 hours). The solvent was removed underreduced pressure. The residue was diluted with CH₂Cl₂ (30 mL) and washedwith 1 N HCl (30 mL×3). The organic phase was dried over anhydrousNa₂SO₄, and concentrated to give the crude product as yellow solid (150mg; yield: 62.6%). The structure was confirmed by ¹H-NMR & LC-MS.

b. Preparation of Compound C0071-2

To the solution of compound C0071-1 (145 mg, 0.376 mmol) in ethanol (3mL) was added 2-aminoethanol (0.44 mL) and p-toluenesulfonic acidmonohydrate (2 mg, 0.01 mmol). The mixture was stirred overnight (about18 hours) at room temperature. The solvent was removed under reducedpressure. The residue was diluted with CH₂Cl₂ (30 mL) and washed withsaturated aqueous NaHCO₃ (30 mL×6). The organic phase was dried overanhydrous Na₂SO₄, then concentrated to give compound C0071-2 as yellowoil (140 mg; yield: 86%). The structure was confirmed by ¹H NMR & LC-MS.

Preparation of Compound C0077

a. Preparation of Compound C0077-1

Piperidin-4-one hydrochloride hydrate (153.5 mg, 1 mmol) in pyridine (2mL) was treated with 4-bromo-3-(trifluoromethyl)benzene-1-sulfonylchloride (323.5 mg, 1 mmol). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. To the residue was added CH₂Cl₂ (50 mL), then the solution waswashed with 1N HCl (20 mL×3), dried over Na₂SO₄ and concentrated to givethe title product as white solid (216 mg; yield: 56%).

b. Preparation of Compound C0077-2

p-Toluenesulfonic acid monohydrate (3.1 mg) and 2-aminoethanol (0.62 mL,10.7 mmol) were added to a solution of compound C0077-1 (207 mg, 0.54mmol) in ethanol (5 mL). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. To the residue was added CH₂Cl₂ (70 mL) and washed withsaturated NaHCO₃ (25 mL×4), then dried over Na₂SO₄ and concentrated togive the product as yellow oil (210 mg, yield: 90.7%).

Preparation of Compound C0078

a. Preparation of Compound C0078-1

To a solution of piperidin-4-one hydrochloride hydrate (169 mg, 1.1mmol) in pyridine (2 mL) was added 4-bromo-2-fluorobenzene-1-sulfonylchloride (300 mg, 1.1 mmol). The mixture was stirred at room temperatureovernight (about 18 hours). The solvent was removed under reducedpressure. The residue was diluted with CH₂Cl₂ (30 mL) and washed with 1NHCl (30 mL×3). The organic phase was dried over anhydrous Na₂SO₄, andconcentrated to give the crude product as white solid (230 mg, yield:62.4%).

b. Preparation of Compound C0078-2

2-Aminoethanol (0.78 mL, 13.39 mmol) and p-toluenesulfonic acidmonohydrate (3.8 mg, 0.02 mmol) was added to the solution of compoundC0078-1 (225 mg, 0.67 mmol) in ethanol (7 mL). The mixture was stirredovernight (about 18 hours) at room temperature. Then the solvent wasremoved under reduced pressure. The residue was diluted with CH₂Cl₂ (30mL) and washed with saturated aqueous NaHCO₃ (30 mL×6). The organicphase was dried over anhydrous Na₂SO₄, and concentrated to give compoundC0078-2 as yellow oil (260 mg)

Preparation of Compound C0079M-7

a. Preparation of Compound C0079M-5

Compound C0079M-4 2HCl (311 mg, 1.44 mmol) was dissolved in water (20ml) and was added saturated aqueous NaHCO₃ to pH=7 and washed withdichloromethane. The water phase was concentrated and the residue wasdissolved in ethanol. The mixture was filtered and the filtrate wasconcentrated to afford 270 mg of compound C0079M-4. A mixture ofC0079M-4 and t-butyl-4-oxopiperidine-1-carboxylate (344 mg, 1.73 mmol)in ethanol (5 ml) was stirred overnight (about 18 hours) at roomtemperature.

b. Preparation of Compound C0079M-6

The mixture of compound C0079M-5 (967 mg, 2.48 mmol) and4-acetylbenzene-1-sulfonyl chloride (597 mg, 2.73 mmol) in pyridine (10ml) was stirred overnight at room temperature. TLC suggested thereaction complete. The mixture was concentrated to remove the solvent.The residue was dissolved in dichloromethane and was washed with 0.1 NHCl and brine, dried, concentrated to afford 854 mg of crude product.The crude product was purified via column chromatography(CH₂Cl₂—CH₂Cl₂/CH₃OH=10/1) to afford a product (440 mg, yield: 40%).

Preparation of Compound C0080M-6

a. Preparation of Compound C0080M-1

Boc-D-Lys-OH (2.46 g, 9.4 mmol) was suspended in CH₂OH (80 mL), and 40%aq HCHO (1.6 mL, 19 mmol) was added, followed by 10% Pd/C (200 mg). Thereaction mixture was stirred under H₂ for 2 days. Then the catalyst wasfiltered off, and the filtrate was concentrated to dryness. Diethylether (100 mL) was added to the resulting oil, and stirring wascontinued to obtain a white solid, which was filtered off and washedwith diethyl ether (2.6 g, yield: 100%, confirmed by ¹H NMR).

b. Preparation of Compound C0080M-2

C0080-1 (375 mg, 1.44 mmol) was dissolved in HCl/CH₂OH (8 mL), and thereaction mixture was stirred at reflux for 3 hours. The mixture wasconcentrated in vacuo to remove the solvent to obtain the HCl salt ofthe title product as white solid (354 mg, yield: 99.5%, confirmed by ¹HNMR). Because the yield was low, the aqueous part was concentrated invacuo to obtain yellow oil (57 mg, yield: 55.8%).

c. Preparation of Compound C0080M-3

C0080-2 (354 mg, 1.43 mmol) was suspended in THF (10 mL) and LiAH₄ (326mg, 8.60 mmol) was added slowly at 0° C., and the reaction mixture wasstirred at 0° C. to room temperature for 1.5 hours. Then saturatedNa₂SO₄ was added to quench the reaction and the mixture was filtered,then the organic layer was concentrated in vacuo to obtain the titleproduct as yellow oil (150 mg, yield: 71.8%, confirmed by ¹H NMR).

d. Preparation of Compound C0080-4

To a solution of N-Boc-piperidin-4-one (150 mg, 0.75 mmol) in ethanol (2mL) was added compound C0080-3 (120 mg, 0.75 mmol). The mixture wasstirred at room temperature overnight (about 18 hours). Then the solventwas removed under reduced pressure. To the residue was added CH₂Cl₂ (70mL) and washed with saturated NaHCO₃ (25 mL×4), then dried over Na₂SO₄and concentrated to give the product as yellow oil (234 mg, yield:91.5%).

e. Preparation of Compound C0080-5

4-Acetylbenzenesulfonyl chloride (180 mg, 0.82 mmol) was added to asolution of compound C0080-4 (234 mg, 0.69 mmol) in pyridine (2 mL). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. To the residue was addedCH₂Cl₂ (70 mL) and washed with saturated NaHCO₃ (25 mL×4), dried overNa₂SO₄ and concentrated to give the crude product. The crude product waspurified by chromatograph with CH₂Cl₂:CH₂OH=50:1 to 3:1 to obtain thetitle product as yellow oil (80 mg, yield: 22.2%).

f. Preparation of Compound C0080-6

To a solution of compound C0080-5 (80 mg, 0.15 mmol) in CH₂Cl₂ (3 mL)was added CF₃COOH (0.2 mL). The mixture was stirred at room temperaturefor 2 hours. CH₂Cl₂ was added and the mixture was washed with saturatedNa₂CO₃. The organic layer was dried over Na₂SO₄, concentrated in vacuoto obtain the product as yellow oil (67 mg)

Preparation of Compound C0081M-7

a. Preparation of Compound C0081M-1

To a stirred solution of L-Boc-Ser-OMe (6.85 g, 31.2 mmol) and imidazole(5.3 g, 78 mmol) in dimethylformamide (60 ml) was added dropwiset-butyldimethylsilyl chloride (7.05 g, 46.8 mmol). The reaction wasstirred at 50° C. overnight (about 18 hours) after which the solvent wasremoved in vacuo. The residue was dissolved in dichloromethane andsuccessively washed with a 1 N HCl solution and brine. The organic layerwas dried over Na₂SO₄ and the solvent was evaporated in vacuo to givethe crude product. The crude reaction product was subjected to columnchromatography (petroleum ether:dichloromethane=1:1) to obtain the titleproduct as light yellow oil (7.4 g; yield: 71.2%).

b. Preparation of Compound C0081M-2

DIBAL-H was added to a solution of C0081M-1 (13.8 g, 41.4 mmol) in drytoluene (100 ml) (1.2 M in toluene, 41.4 ml, 50 mmol) in an argonatmosphere at such a rate that the temperature didn't rise above 70° C.After stirring for 2 hours, HCl/CH₃OH solution was added carefully. Thereaction mixture was poured into 10% aqueous HCl solution. The organiclayer was separated and washed with brine. The organic layer was driedover Na₂SO₄ and concentrated to get the crude product. The crude productwas purified with column chromatography (petroleum ether:ethylacetate=40:1 to 10:1) to obtain the title product as a colorless liquid(8.24 g, yield: 66%).

c. Preparation of Compound C0081M-3

To a solution of C0081M-2 (8.24 g, 27 mmol) in methanol (75 ml) wasadded NaOAc (2.16 g, 28 mmol) and dimethylamine hydrochloride (2.756 g,33.8 mmol). The mixture was stirred at room temperature for 1 hour, thenNaCNBH₃ (1.7 g, 27 mmol) was added. The mixture was allowed to stir atroom temperature overnight (about 18 hours). Water was added and thesolution was extracted with CH₂Cl₂. The combined organic layers wasdried over Na₂SO₄ and concentrated to get the crude product. The crudeproduct was purified with column chromatography (petroleum ether:ethylacetate=40:1 to 1:1) to obtain the title product as a colorless liquid(2.512 g, yield: 28%).

Preparation of Compound C0083M

This compound is prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0086M

a. Preparation of Compound C0086M-1

To a stirred solution of glycinamide hydrochloride (276 mg, 2.5 mmol)and triethylamine (0.35 mL, 2.5 mmol) in methanol (10 mL) was addedN-Boc-piperidin-4-one (500 mg, 2.5 mmol). The reaction was stirred underreflux overnight (about 18 hours) after which the solvent was removed invacuo. The residue was purified by column chromatography to obtain thetitle product as white solid (500 mg; yield: 78%).

b. Preparation of Compound C0086M-2

4-Acetylbenzene-1-sulfonyl chloride (258 mg, 1.18 mmol) was added to asolution of compound C0086M-1 (250 mg, 0.98 mmol) in pyridine (10 mL).The mixture was stirred at 60° C. (yield: 46%) overnight (about 18hours). Then the solvent was removed under reduced pressure. The residuewas purified by column chromatography to obtain the title product asyellow solid (250 mg; yield: 58%)

c. Preparation of Compound C0086M

To a solution of compound C0086M-2 (150 mg, 0.343 mmol) in 5 mLdichloromethane was add 0.5 mL CF₃COOH, and the mixture was stirred for2 hours at room temperature. To the mixture was added 30 mLdichloromethane, and the composition formed was washed with saturatedsodium carbonate solution (30 mL×3). The organic layer was dried andconcentrated to get 90 mg of the crude product as light yellow solid.The crude product was purified by column chromatography to obtain thetitle product as little yellow solid (50 mg; yield: 43.2%).

¹H NMR (400 MHz, DMSO): 9.49 (s, 1H); 8.14 (d, J=8.4 Hz, 2H); 8.04 (d,J=8.4 Hz, 2H); 3.93 (s, 2H); 2.87 (m, 2H); 2.72-2.59 (m, 2H); 2.65 (s,3H), 2.33 (td, J=12.4, 4.4 Hz, 2H); 1.51 (d, J=12.0 Hz, 2H); LCMS (ESI)calcd for C₁₅H₁₉N₃O₄S (m/z): 337.39. found: 338.4[M+1]⁺.

Preparation of Compound C0087M

2-Hydroxyacetamide (210 mg, 2.8 mmol) and toluenesulfonic acidmonohydrate (3.0 mg) was added to a solution of C0066-1 (200 mg, 0.7mmol) in dimethylformamide (10 mL). The mixture was stirred at 120° C.overnight (about 18 hours). The solvent was removed under reducedpressure. The residue was diluted with dichloromethane and a yellowsolid formed. The mixture was filtered and the filtrate was concentratedand purified by column chromatography (dichloromethane:methanol=100:1 to30:1) to obtain the impure product as yellow solid (30 mg, yield: 13%).The impure product was recrystallized with chloroform and n-hexane toobtain the title product as yellow solid (20 mg, Yield: 8%).

¹H NMR (400 MHz, CDCl₃): 8.11 (d, J=9.2 Hz, 2H); 7.87 (d, J=8.4 Hz, 2H);6.71 (s, 1H); 4.15 (s, 2H); 3.65 (m, 2H); 2.83-2.77 (m, 2H); 2.67 (s,3H); 1.97-1.88 (m, 4H); MS (ESI) calcd for C₁₅H₁₈N₂O₅S (m/z): 338.38.found: 339.4 [M+1]⁺.

Preparation of Compound C0088M

a. Preparation of Compound C0088M-1

2-aminoethanol (0.13 mL, 2.1 mmol) was added to a solution of1-methylpiperidin-4-one (227 mg, 2.0 mmol) in ethanol (4 mL). Thereaction was stirred under reflux for 2 hours. The solvent was removedin vacuo to obtain the crude product C0088M-1 as yellow oil. The crudeproduct was used for the next step directly with out purification.

b. Preparation of Compound C0088M

To a solution of compound C0088M-1 in pyridine (5 mL) was added4-methoxybenzene-1-sulfonyl chloride (207 mg, 1.0 mmol). The mixture wasstirred at room temperature overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography to obtain the crude product. The crude product waspurified by preparative HPLC to obtain the title product as yellow oil(35 mg; yield: 10.7%; confirmed by ¹H NMR and LC-MS, purity: 98.1% byHPLC).

¹H NMR (400 MHz, CDCl₃): 7.80 (d, J=8.4 Hz, 2H); 6.96 (d, J=8.4 Hz, 2H);3.94 (t, J=6.0 Hz, 2H); 3.87 (s, 3H); 3.50 (t, J=6.0 Hz, 2H); 2.74 (m,2H); 2.53 (td, J=12.8, 4.4 Hz, 2H); 2.27 (s, 3H); 2.19 (t, J=11.6 Hz,2H); 1.57 (d, J=12.4 Hz, 2H); LCMS (ESI) calcd for C₁₅H₂₂N₂O₄S (m/z):326.41. found: 327.3[M+1]⁺.

Preparation of Compound C0089M

a. Preparation of Compound C0089M-1

To a solution of 1-ethylpiperidin-4-one (254 mg, 2.0 mmol) in ethanol(4.0 mL) was added 2-aminoethanol (0.13 mL, 2.1 mmol). The reaction wasrefluxed for 3 hours. The solvent was removed in vacuo to obtain thecrude product of C0094M-1 as a red oil. The crude product was used forthe next step directly with out purification.

b. Preparation of Compound C0089M

The crude C0089M-1 was dissolved in pyridine (2.5 mL) and added4-methoxybenzenesulfonyl chloride (207 mg, 1.0 mmol). The mixture wasstirred at room temperature overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography to give the crude product as yellow solid (80 mg; yield:24%). The crude product was purified by preparative HPLC to obtain thetitle product as white solid (40 mg; yield: 11.5%, confirmed by ¹H-NMRand LC-MS, purity: 99% by HPLC).

¹H NMR (400 MHz, CDCl₃): 7.80 (d, J=9.2 Hz, 2H); 6.96 (d, J=9.2 Hz, 2H);3.93 (t, J=6.0 Hz, 2H); 3.86 (s, 3H); 3.50 (t, J=6.0 Hz, 2H); 2.86-2.82(m, 2H); 2.52-2.49 (m, 2H); 2.42 (q, J=7.2 Hz, 2H); 2.16 (t, J=11.2 Hz,2H); 1.60-1.56 (m, 2H); 1.07 (t, J=7.2 Hz, 3H); MS (ESI) calcd forC₁₆H₂₄N₂O₄S (m/z): 340.44. found: 341.4 [M+1]⁺.

Preparation of Compound C0090M

C0027 (200 mg, 0.64 mmol) was dissolved in acetone (20 mL), and(bromomethyl)cyclopropane (86 mg, 0.64 mmol) was added, followed byK₂CO₃ (265 mg, 1.92 mmol). The reaction mixture was stirred at refluxfor 7 hours and then stirred at room temperature overnight (about 18hours). The mixture was filtered and concentrated in vacuo to obtain thecrude product. The crude product was purified by column chromatographyeluted with ethyl acetate to obtain 220 mg of the title product as lightyellow oil (yield: 93.9%). The structure was confirmed by ¹H NMR &LC-MS. Purity: 99.6% by HPLC.

¹H NMR (400 MHz, CDCl₃): 7.81 (d, J=8.4 Hz, 2H); 6.96 (d, J=8.4 Hz, 2H);3.93 (t, J=6.0 Hz, 2H); 3.87 (s, 3H); 3.50 (t, J=6.0 Hz, 2H); 2.97 (m,2H); 2.52 (dt, J=4.4, 10.2 Hz, 2H); 2.26-2.17 (m, 4H); 1.58 (m, 2H);0.92-0.80 (m, 1H); 0.52-0.48 (m, 2H); 0.09-0.05 (m, 2H); MS (ESI) calcdfor C₁₈H₂₆N₂O₄S (m/z): 366.16. found; 367.2 [M+1]⁺.

Preparation of Compound C0091M

C0027 (50 mg, 0.16 mmol) was dissolved in acetone (5 mL), and3-bromoprop-1-ene (20 mg, 0.16 mmol) was added, followed by K₂CO₃ (66mg, 0.48 mmol). The reaction mixture was stirred at reflux for 7 hours,and then stirred at room temperature overnight (about 18 hours). Themixture was filtered and concentrated in vacuo to obtain the crudeproduct. The crude product was purified by column chromatography withethyl acetate to obtain 30 mg of the title product as colorless oil (30mg, yield: 53.3%). The structure was confirmed by ¹H NMR & LC-MS,purity: 99.9% by HPLC.

¹H NMR (400 MHz, CDCl₃): 7.79 (d, J=8.4 Hz, 2H); 6.95 (d, J=8.48 Hz,2H); 5.92-5.84 (m, 1H); 5.18-5.10 (m, 2H); 3.92 (t, J=6.4 Hz, 2H); 3.86(s, 3H); 3.48 (t, J=6.4 Hz, 2H); 2.99 (d, J=6.0 Hz, 2H); 2.82 (dt,J=2.8, 11.2 Hz, 2H); 2.50 (dt, J=4.8, 12.8 Hz, 2H); 2.16 (dt, J=2.0,13.6 Hz, 2H); 1.57 (d, J=11.2 Hz, 2H); MS (ESI) calcd for C₁₇H₂₄N₂O₄S(m/z): 352.15. found: 353.4 [M+1]⁺.

Preparation of Compound C0092M

This compound is prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0093M

a. Preparation of Compound C0088M-1

2-Aminoethanol (0.13 mL, 2.1 mmol) was added to a solution of1-methylpiperidin-4-one (227 mg, 2.0 mmol) in ethanol (4 mL). Thereaction was stirred under reflux for 2 hours. The solvent was removedin vacuo to obtain the crude product of C0088M-1 as yellow oil. Thecrude product was used for the next step directly with out purification.

b. Preparation of Compound C0093M

To a solution of compound C0088M-1 in pyridine (5 mL) was added4-methoxybenzene-1-sulfonyl chloride (219 mg, 1.0 mmol). The mixture wasstirred at room temperature overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography to obtain the crude product. The crude product waspurified by preparative HPLC to obtain the title product as yellow oil(20 mg; yield: 6.0%; confirmed by ¹H-NMR and LC-MS, purity: 99% byHPLC).

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.0 Hz, 2H); 7.97 (d, J=8.0 Hz, 2H);3.97 (t, J=6.0 Hz, 2H); 3.56 (t, J=6.0 Hz, 2H); 2.75 (m, 2H); 2.66 (s,3H); 2.53 (td, J=12.8, 4.4 Hz, 2H); 2.28 (s, 3H); 2.20 (t, J=11.6 Hz,2H); 1.56 (d, J=12.4 Hz, 2H); LCMS (ESI) calcd for C₁₆H₂₂N₂O₄S (m/z):338.42. found: 339.5 [M+1]⁺.

Preparation of Compound C0094M

a. Preparation of Compound C0094M-1

To a solution of 1-ethylpiperidin-4-one (254 mg, 2.0 mmol) in ethanol(4.0 mL) was added 2-aminoethanol (0.13 mL, 2.1 mmol). The reaction wasrefluxed for 3 hours. Then the solvent was removed in vacuo to obtainthe crude product of C0094M-1 as a red oil.

b. Preparation of Compound C0094M

The crude C0094-1 was dissolved in pyridine (2.5 mL) and added4-acetylbenzene-1-sulfonyl chloride (218 mg, 1.0 mmol). The mixture wasstirred at room temperature overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography to give the crude product as yellow solid (90 mg; yield:26%). The crude product was purified by Pre-HPLC to obtain the titleproduct as white solid (50 mg; yield: 14%, confirmed by ¹H NMR andLC-MS, purity: 99% by HPLC).

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.4 Hz, 2H); 7.97 (d, J=8.8 Hz, 2H);3.97 (t, J=6.4 Hz, 2H); 3.57 (t, J=6.0 Hz, 2H); 2.87-2.83 (m, 2H); 2.65(s, 3H); 2.52-2.48 (m, 2H); 2.44 (q, J=7.2 Hz, 2H); 2.20-2.17 (m, 2H);1.57 (d, J=10.8 Hz, 2H); 1.08 (t, J=7.2 Hz, 3H); MS (ESI) calcd forC₁₇H₂₄N₂O₄S (m/z): 352.45. found: 353.4 [M+1]⁺.

Preparation of Compound C0095M

To the solution of compound C0095M-2 (260 mg, 1.33 mmol) in pyridine(2.5 mL) was added 4-acetylbenzene-1-sulfonyl chloride (348 mg, 1.59mmol). The mixture was stirred overnight (about 18 hours) at roomtemperature. The reaction mixture was concentrated in vacuo to removethe pyridine. The residue was dissolved into dichloromethane and washedwith saturated NaHCO₃. Then, the organic layer was dried over Na₂SO₄ andconcentrated in vacuo to obtain the crude product. The crude product waspurified by column chromatography to obtain 464 mg of C0095M. Theproduct was purified by preparative HPLC to obtain the title product(240 mg, yield: 47.8%).

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.4 Hz, 2H); 7.97 (d, J=8.4 Hz, 2H);3.96 (t, J=6.0 Hz, 2H); 3.56 (t, J=6.0 Hz, 2H); 2.97 (dt, J=2.4, 11.2Hz, 2H); 2.65 (s, 3H); 2.51 (dt, J=4.8, 12.8 Hz, 2H); 2.25 (d, J=6.8 Hz,2H); 2.22-2.15 (m, 2H); 1.56 (dd, J=2.0, 13.2 Hz, 2H); 0.87-0.83 (m,1H); 0.52-0.48 (m, 2H); 0.09-0.05 (m, 2H); MS (ESI) calcd forC₁₉H₂₆N₂O₄S (m/z): 378.16. found: 379.4 [M+1]⁺.

Preparation of Compound C0096M

C0046 (60 mg, 0.185 mmol) was dissolved in acetone (5 mL), and3-bromoprop-1-ene (22 mg, 0.185 mmol) was added, followed by K₂CO₃ (77mg, 0.556 mmol). The reaction mixture was stirred at reflux for 4 hours,and then stirred at room temperature overnight (about 18 hours). Themixture was filtered and concentrated in vacuo to obtain the crudeproduct. The crude product was purified by column chromatography withethyl acetate to obtain 43 mg of the title product as white solid (43mg, yield: 63.9%)

¹H NMR (400 MHz, CDCl₃): 8.06 (d, J=9.2 Hz, 2H); 7.96 (d, J=9.2 Hz, 2H);5.90-5.83 (m, 1H); 5.18-5.11 (m, 2H); 3.96 (t, J=6.4 Hz, 2H); 3.55 (t,J=6.4 Hz, 2H); 2.99 (d, J=6.4 Hz, 2H); 2.82 (dt, J=2.0, 11.2 Hz, 2H);2.65 (s, 3H); 2.50 (dt, J=4.8, 13.6 Hz, 2H); 2.18 (dt, J=2.4, 9.6 Hz,2H); 1.56 (dd, J=2.4, 10.8 Hz, 2H); MS (ESI) calcd for C₁₈H₂₄N₂O₄S(m/z): 364.15. found: 365.3 [M+1]⁺.

Preparation of Compound C0097M

This compound is prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0099M

Prepared using procedures illustrated elsewhere herein.

¹H NMR (400 MHz, CDCl₃): 8.06 (d, J=8.4 Hz, 2H); 7.96 (d, J=8.4 Hz, 2H);4.18 (q, J=6.8 Hz, 2H); 3.96 (t, J=6.4 Hz, 2H); 3.56 (t, J=6.8 Hz, 2H);3.21 (s, 2H); 2.85 (dd, J=1.2, 8.8 Hz, 2H); 2.66 (s, 3H); 2.57-2.45 (m,4H); 1.54 (d, J=11.2 Hz, 2H); 1.27 (t, J=6.8 Hz, 3H); MS (ESI) calcd forC₁₉H₂₆N₂O₆S (m/z): 410.15. found: 411.4 [M+1]⁺.

Preparation of Compound C0100M

Prepared by hydrolysis of compound C0099M.

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.4 Hz, 2H); 7.91 (d, J=8.4 Hz, 2H);3.91 (t, J=6.4 Hz, 2H); 3.47 (t, J=6.4 Hz, 2H); 3.39 (m, 4H); 2.94 (br,m, 2H); 2.63 (dt, J=4.8, 14.4 Hz, 2H); 2.55 (s, 3H); 1.73 (d, J=12.8 Hz,2H); MS (ESI) calcd for C₁₇H₂₂N₂O₆S (m/z): 382.12. found: 383.2 [M+1]⁺.

Preparation of Compound C0101M

Prepared using procedures illustrated elsewhere herein.

¹H NMR (400 MHz, CDCl₃): 8.06 (d, J=8.0 Hz, 2H); 7.95 (d, J=8.0 Hz, 2H);3.96 (t, J=6.4 Hz, 2H); 3.56 (t, J=6.4 Hz, 2H); 2.81 (dd, J=2.0, 9.2 Hz,2H); 2.66 (s, 3H); 2.48 (dt, J=2.0, 8.4 Hz, 2H); 2.32 (d, J=7.2 Hz, 2H);2.17 (dt, J=1.6, 13.2 Hz, 2H); 1.55 (d, J=11.2 Hz, 2H); 1.47-1.43 (m,2H); 1.34-1.28 (m, 2H); 0.92 (t, J=7.2 Hz, 3H); MS (ESI) calcd forC₁₉H₂₈N₂O₄S (m/z): 380.18. found: 381.4 [M+1]⁺.

Preparation of Compound C0102M

Prepared using procedures illustrated elsewhere herein.

¹H NMR (400 MHz, CDCl₃): 7.80 (d, J=8.0 Hz, 2H); 6.96 (d, J=8.0 Hz, 2H);3.93 (t, J=6.0 Hz, 2H); 3.86 (s, 3H); 3.49 (t, J=6.4 Hz, 2H); 2.81 (dt,J=2.0, 10.8 Hz, 2H); 2.49 (dt, J=4.8, 12.8 Hz, 2H); 2.33-2.29 (m, 2H);2.16 (dt, J=2.8, 13.2 Hz, 2H); 1.57 (d, J=10.4 Hz, 2H); 1.47-1.42 (m,2H); 1.34-1.28 (m, 2H); 0.91 (t, J=7.6 Hz, 3H); MS (ESI) calcd forC₁₈H₂₈N₂O₄S (m/z): 368.18. found: 369.3 [M+1]⁺.

Preparation of Compound C0104M

a. Preparation of Compound C0104M-1

A mixture of 2-aminoacetamide hydrochloride (2.0 g, 18 mmol),triethylamine (3.7 mL) and benzaldehyde (2.3 g, 22 mmol) in methanol (40mL) was stirred overnight (about 18 hours) at room temperature. To thereaction mixture NaBH₄ (1.0 g) was added slowly. The reaction mixturewas stirred for 1.5 hours until thin-layer chromatography showed thereaction complete. The solid was filtered. The filtrate was concentratedto remove the solvent. The residue was dissolved in water and ethylacetate (EA). The mixture was extracted with EA and the organic layerswere combined and dried and concentrated. Ether was added to the residueether when the solid was appeared. The solid was dried to obtain thefinal product (1.3 g, Yield: 45%).

b. Preparation of Compound C0104M-1

C0104M-1 (100 mg, 0.61 mmol) was dissolved in methanol (5 mL) and1-methyl-4-piperidone (69 mg, 0.61 mmol) was added, followed bytriethylamine 0.085 mL, 0.61 mmol). The reaction mixture was stirred atreflux overnight (about 18 hours). The mixture was concentrated in vacuoto obtain the crude product. The crude product was purified bychromatography with CH₂Cl₂:CH₂OH=50:1 to 10:1 to obtain 75 mg of thetitle product as white solid (yield: 48%).

Preparation of Compound C0105M

C0103M (338 mg, 2.0 mmol) was dissolved in dimethylformamide (10 mL) andNaH (840 mg, 20.0 mmol) was added. The reaction mixture was stirred atroom temperature for 1 hour. Then benzylchloride (253 mg, 2.0 mmol) wasadded, and the mixture was stirred at room temperature overnight (about18 hours). H₂O was added to quench the reaction and the mixture wasextracted with dichloromethane (DCM) (3×). The organic layer was washedwith 1M HCl (3×) and the aqueous layer was washed with DCM, alkalizedwith 1M NaOH to pH 8, and extracted with DCM (3×). The organic layer waswashed with saturated NaCl (1×), dried over Na₂SO₄, and thenconcentrated in vacuo to obtain the crude product. The crude product waspurified by chromatography with DCM:actone=4:1 to 2:1 and DCM:CH₃OH=10:1to 6:1 to obtain 110 mg of the product as colorless oil (110 mg, yield:21.2%).

Preparation of Compound C0106M-1

a. Preparation of Compound C0106M-1

1-Methyl-4-piperidone (200 mg, 1.77 mmol) was dissolved in ethanol (2mL), and ethanol amine (97 mg, 1.60 mmol) was added. The reactionmixture was stirred at room temperature overnight (about 18 hours). Thereaction mixture was concentrated in vacuo to obtain the crude product.The crude product was used by next step without any purification (184mg, yield: 66.6%).

a. Preparation of Compound C0106M

To the solution of compound C0106M-1 (184 mg, 1.18 mmol) in pyridine(2.5 mL) was added 4-methylsulfonylbenzenesulfonyl chloride (360 mg,1.42 mmol). The mixture was stirred overnight (about 18 hours) at roomtemperature. The precipitated was filtrated to obtain the crude product.The crude product was purified by column chromatography to obtain 170 mgof the title product as light yellow solid (170 mg, yield: 38.5%).

¹H NMR (400 MHz, CD₃OD): 8.09-8.02 (m, 4H); 3.92 (t, J=6.4 Hz, 2H); 3.48(t, J=6.8 Hz, 2H); 3.19-3.15 (m, 2H); 3.08 (s, 3H); 2.91-2.83 (m, 2H);2.59 (s, 3H), 2.58-2.51 (m, 2H); 1.75 (d, J=14.8 Hz, 2H); calcd forC₁₅H₂₂N₂O₅S₂ (m/z): 374.1. found: 375.2 [M+1]⁺.

Preparation of Compound C0108M

a. Preparation of Compound C0108M-1

To a solution of N-methyl-piperidone (227 mg, 2.1 mmol) in ethanol (2mL) was added compound C0065-3 (213 mg, 1.8 mmol). The mixture wasstirred overnight (about 18 hours) at room temperature. The solvent wasremoved under reduced pressure to give the crude product as yellow oil.The crude product was used for next step without any furtherpurification (440 mg; yield: 100%).

b. Preparation of Compound C0108M

4-Acetylbenzene-1-sulfonyl chloride (473 mg, 2.2 mmol) was added to asolution of compound C0108M-1 (384 mg, 1.8 mmol) in pyridine (2 mL). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. The residue was diluted withdichloromethane (100 mL) and washed with saturated sodium carbonatesolution (50 mL×3). The organic layer was dried and concentrated to getthe crude product. The crude product was purified by columnchromatography (dichloromethane:acetone=1:1) to obtain the titlecompound as yellow solid. 100 mg of crude compound C0108M was purifiedby preparative thin-layer chromatography (CH₂Cl₂:CH₂OH:MeOH=8:1) to getthe compound as little yellow solid (35 mg; yield: 5%).

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.4 Hz, 2H); 7.96 (d, J=8.4 Hz, 2H);4.24-4.20 (m, 1H); 3.75 (dd, J=8.4, 5.6 Hz, 1H); 3.11 (t, J=8.8 Hz, 1H);2.74 (m, 2H); 2.66 (s, 3H); 2.61 (dd, J=12.8, 4.4 Hz, 1H); 2.46 (d,J=5.6 Hz, 2H); 2.40 (dd, J=13.2, 4.4 Hz, 1H); 2.27 (s, 3H), 2.26 (s,3H), 2.24-2.18 (m, 5H); 1.72 (d, J=10.8 Hz, 1H); 1.41 (d, J=10.8 Hz,1H); LCMS (ESI) calcd for C₁₉H₂₉N₃O₄S (m/z): 395.52. found: 396.4[M+1]+.

Preparation of Compound C0109M

a. Preparation of Compound C0109M-1

To a solution of compound C0084M-2 (617 mg, 4.0 mmol) in methanol (15mL) was added 1-methylpiperidin-4-one (227 mg, 2.0 mmol). The mixturewas stirred at 70° C. overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography (dichloromethane:methanol=2:1) to obtain the titlecompound as yellow solid (130 mg; yield: 26%).

Preparation of Compound C0111M

This compound is prepared using procedures illustrated elsewhere herein.

Preparation of Compound C0114M

a. Preparation of Compound C0115M-1

A stirred solution of glycinamide hydrochloride (1.65 g, 15 mmol) intriethylamine (2.1 mL) and methanol (50 mL) was treated withN-Boc-piperidin-4-one (3.0 g, 15 mmol). The reaction was stirred underreflux overnight (about 18 hours) after which the solvent was removed invacuo. The residue was purified by column chromatography (CH₂Cl₂ toCH₂Cl₂:CH₂OH=10:1) to obtain the title product as white solid (3.04 g;yield: 79%). The structure was confirmed by ¹H NMR.

b. Preparation of Compound C0114M-1

To a solution of C0115M-1 (100 mg, 0.4 mmol) in dimethylformamide (DMF)(2.5 mL), was added 64 mg of NaH as 60% dispersion in mineral oil. Themixture was stirred at room temperature for 1 hour. Then a solution ofdichloromethane (55 mg, 0.32 mmol) in 0.5 mL of absolute DMF was addeddropwise. The mixture was stirred overnight (about 18 hours). Water wasadded and the solution was extracted with dichloromethane. The organiclayer was washed with water, dried over anhydrous Na₂SO₄ andconcentrated. The residue was purified by preparative thin-layerchromatography (CH₂Cl₂:CH₂OH:MeOH=30:1) to obtain the crude product asyellow oil (target M+1=345) (30 mg). Judged by ¹H NMR & LC-MS, theproduct was a mixture of the following 2 compounds.

The mixture was purified by preparative thin-layer chromatography(dichloromethane:acetone=4:1) to obtain the title product as yellow oil(9 mg).

c. Preparation of Compound C0114M

To the solution of C0114M (200 mg, 0.58 mmol) in 3.0 mL ofdichloromethane was add 0.3 mL of CF₃COOH and the mixture was stirredfor 1 hour at room temperature. Then CH₃CH₂OH/NH₃ was added and thesolvent was removed under reduced pressure. 10 mL of dichloromethane wasadded and white solid formed. The mixture was filtered and the filtratewas concentrated under reduced pressure. The residue was purified withcolumn chromatography using a stationary phase of aluminum oxide(CH₂Cl₂:CH₃OH=100:1 to 30:1) to obtain the title product as colorlessoil (30 mg, yield: 21%).

¹H NMR (400 MHz, CDCl₃): 7.32-7.23 (m, 5H); 4.46 (s, 2H); 3.54 (s, 2H);2.99-2.89 (m, 4H); 2.02 (s, 2H); 1.86-1.79 (m, 2H); 1.43-1.40 (m, 2H);MS (ESI) calcd for C₁₄H₁₉N₃O (m/z): 245.32. found: 246.3 [M+1]+.

Preparation of Compound C0115M

a. Preparation of Compound C0115M-1

N-Boc-piperidin-4-one (3.0 g, 15 mmol) was added to a stirred solutionof glycinamide hydrochloride (1.65 g, 15 mmol) in triethylamine (2.1 mL)and methanol (50 mL). The reaction was stirred under reflux overnight(about 18 hours) after which the solvent was removed in vacuo. Theresidue was purified by column chromatography (CH₂Cl₂ toCH₂Cl₂:CH₂OH=10:1) to obtain the title product as white solid (3.04 g;yield: 79%).

b. Preparation of Compound C0115M-2

To a solution of C0115M-1 (306 mg, 1.2 mmol) in dimethylformamide (5.0mL) was added 1-(bromomethyl)benzene (225 mg, 1.32 mmol). The mixturewas allowed to stir at room temperature overnight (about 18 hours).Water was added and white solid was formed. The mixture was filtered andthe collected solid was washed with water and n-hexane. Then the solidwas dried to obtain the title product as white solid (200 mg, Yield:48%, confirmed by ¹H NMR).

c. Preparation of Compound C0115M

To the solution of C0115M-2 (200 mg, 0.58 mmol) in 3.0 mL ofdichloromethane was add 0.3 mL of CF₃COOH and the mixture was keptstirring for 1 hour at room temperature. The solvent was removed underreduced pressure quickly and CH₂Cl₂OH/NH₃ was added. Thin-layerchromatography suggested there was new compound formed. The solvent wasremoved under reduced pressure and the residue was purified withaluminum oxide chromatography (CH₂Cl₂:CH₂OH=100:1 to 5:1) to obtain the2 batches of product as white solid (40 mg, yield: 28%, HPLC: 90%; 30mg, yield: 21%, HPLC: 82%). The 40 mg of sample C0115 was recrystallizedwith chloroform and n-hexane to obtain the title product as white solid(19 mg, yield 14%). The structure was confirmed by ¹H NMR and LC-MS(target M+1=246).

¹H NMR (400 MHz, CDCl₃): 8.18 (s, 1H); 7.34-7.24 (m, 5H); 3.77 (s, 2H);3.23 (s, 2H); 3.15 (d, J=12.0 Hz, 2H); 2.77 (t, J=12.0 Hz, 2H);1.98-1.91 (m, 2H); 1.73-1.68 (m, 2H); MS (ESI) calcd for C₁₄H₁₉N₃O(m/z): 245.32. found: 246.3 [M+1]⁺.

Preparation of Compound C0116M

a. Preparation of Compound C0116M-1

To a solution of N-Boc-piperidin-4-one (0.3 g, 1.5 mmol) in ethanol (3.0mL) was added ethanolamine (0.3 mL). The mixture was stirred overnight(about 18 hour) at room temperature. The solvent was removed underreduced pressure. The residue was diluted with CH₂Cl₂ and washed withsaturated aqueous Na₂CO₃. The organic phase was dried over anhydrousNa₂SO₄, concentrated to obtain the title product as yellow oil (327 mg,yield: 90%)

b. Preparation of Compound C0116M-2

4-(Methylsulfonyl)-benzene-1-sulfonyl chloride (305 mg, 1.2 mmol) wasadded to a solution of C0116-1 (242 mg, 1.0 mmol) in pyridine (2.0 mL).The mixture was stirred at room temperature overnight (about 18 hours).The solvent was removed under reduced pressure. The residue was dilutedwith CH₂Cl₂ and washed with saturated NaHCO₃. The organic phase wasdried over anhydrous Na₂SO₄ and concentrated to give the crude productas yellow solid, which was purified with column chromatography (CH₂Cl₂to CH₂Cl₂:CH₂OH=50:1) to give the title compound as a white solid (270mg, yield: 58%, ¹H NMR confirmed, confirmed by ¹H NMR and LC/MS (targetM+23=483).

c. Preparation of Compound C0116M

To the solution of C0016M-2 (270 mg, 0.58 mmol) in 3.0 mLdichloromethane (DCM) was add 0.3 mL CF₃COOH and the mixture was keptstirring for 1 hour at room temperature. The mixture was added DCM andwashed with saturated sodium carbonate solution. The organic layer wasdried and concentrated to get the crude product as yellow gum. The crudeproduct was purified by column chromatography (CH₂Cl₂:CH₂OH-100:1 to10:1) to obtain the product as yellow solid (70 mg, yield: 34%).

¹H NMR (400 MHz, CDCl₃): 8.12-8.07 (m, 4H); 4.01 (t, J=6.0 Hz, 2H); 3.55(t, J=6.0 Hz, 2H); 3.11 (s, 3H); 3.04-3.00 (m, 2H); 2.86-2.79 (m, 2H);2.35-2.28 (m, 2H); 1.63-1.60 (m, 2H); calcd for C₁₄H₂₀N₂O₅S₂ (m/z):360.45. found: 361.2 [M+1]⁺.

Preparation of Compound C0118M

a. Preparation of Compound C0118M-1

To a solution of N-Boc-piperidin-4-one (420 mg, 2.1 mmol) in ethanol (4mL) was added compound C0065-3 (500 mg, 4.2 mmol). The mixture wasstirred overnight (about 18 hours) at room temperature. The solvent wasremoved under reduced pressure. The residue was diluted with CH₂Cl₂ (50mL) and washed with saturated aqueous Na₂CO₃ (30 mL×6). The organicphase was dried over anhydrous Na₂SO₄, and concentrated to give compoundC0118M-1 as yellow oil (600 mg; yield: 95%).

b. Preparation of Compound C0118M-2

4-Acetylbenzene-1-sulfonyl chloride (525 mg, 2.4 mmol) was added to asolution of compound C0118M-1 (600 mg, 2 mmol) in pyridine (2 mL). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. The residue was purified bycolumn chromatography (dichloromethane:acetone=3:1) to obtain the titlecompound as light yellow solid (610 mg; yield: 63.4%).

c. Preparation of Compound C0118M

To the solution of compound C0118M-2 (610 mg, 1.26 mmol) in 10 mLdichloromethane (DCM) was added 0.5 mL CF₃COOH and the mixture wasstirred for 1 hour at room temperature. Then DCM (30 mL) was added tothe mixture, and the resulting mixture washed with saturated sodiumcarbonate solution (30 mL×3). The organic layer was dried andconcentrated to get the crude product. The crude product was purified bycolumn chromatography (CH₂Cl₂:CH₃CH₂OH=10:1 to CH₃OH) to obtain thetitle compound as light yellow solid (60 mg; yield: 12.5%).

¹H NMR (400 MHz, CDCl₃): 8.07 (d, J=8.4 Hz, 2H); 7.96 (d, J=8.4 Hz, 2H);4.27-4.21 (m, 1H); 3.73 (dd, J=8.4, 5.6 Hz, 1H); 3.08 (t, J=8.8 Hz, 1H);3.04-2.94 (m, 2H), 2.83 (t, J=11.6 Hz, 2H); 2.66 (s, 3H); 2.46 (d, J=5.6Hz, 2H); 2.41 (dd, J=12.4, 5.2 Hz, 1H); 2.26 (s, 6H); 2.20 (dd, J=12.8,4.8 Hz, 1H); 1.78 (d, J=13.2 Hz, 1H); 1.45 (d, J=12.8 Hz, 1H); LCMS(ESI) calcd for C₁₈H₂₇N₃O₄S (m/z): 381.49. found: 382.4 [M+1]⁺.

Preparation of Compound C0119M

a. Preparation of Compound C0119M-1

N-Boc-piperidin-4-one (797 mg, 4.0 mmol) was added to a solution ofcompound C0084M-2 (617 mg, 4.0 mmol) in methanol (15 mL). The mixturewas stirred at 70° C. overnight (about 18 hours). The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography (CH₂Cl₂:CH₃OH=8:1) to obtain the title compound as whitesolid (300 mg; yield: 22.4%).

b. Preparation of Compound C0119M-2

4-Acetyl-benzene-1-sulfonyl chloride (149 mg, 0.68 mmol) andtriethylamine (1 mL) was added to a solution of compound C0119M-1 (190mg, 0.57 mmol) in chloroform (5 mL). The mixture was stirred underreflux overnight (about 18 hours). The solvent was removed under reducedpressure. The residue was purified by column chromatography(dichloromethane:acetone=3:1) to obtain the product as white solid (250mg; yield: 84%).

c. Preparation of Compound C0119M

To the solution of compound C0119M-2 (330 mg, 0.63 mmol) in 3.5 mLdichloromethane (DCM) was add CF₃COOH (0.35 mL) and the mixture was keptstirring for 1 hour at room temperature. Then CH₃CH₂OH/NH₃ (20 mL) wasadded to the reaction mixture. The solvent was removed under reducedpressure. The residue was diluted with dichloromethane (20 mL) and thewhite solid was precipitated. TLC showed that the solid was CF₃COONH₄.The mixture was filtered and the filtrate was concentrated. The residuewas purified by column chromatography (CH₂Cl₂:CH₃OH=8:1) to obtain theproduct as white solid (80 mg; yield: 30%).

¹H NMR (400 MHz, CD₃OD): 8.17 (s, 1H); 8.11 (m, 4H); 7.36 (s, 1H); 3.78(dd, J=8.0, 4.0 Hz, 1H); 3.22-3.11 (m, 6H); 2.89 (dd, J=15.2, 4.0 Hz,1H); 2.67 (dd, J=15.2, 8.0 Hz, 1H); 2.56 (s, 3H); 1.89-1.73 (m, 3H);LCMS (ESI) calcd for C₁₉H₂₃N₅O₄S (m/z): 417.48. found: 418.5 [M+1]⁺.

Preparation of Compound C0125M

A mixture of C0104M-1 (350 mg, 2.13 mmol), 1-(cyclopropylmethyl)piperidin-4-one (326 mg, 2.13 mmol) and p-toluenesulfonic acid (10 mg)in methanol (15 mL) was heated to reflux overnight (about 18 hours).Thin-layer chromatography suggested the reaction complete. The solventwas removed. The residue was dissolved in DCM and washed with saturatedaqueous NaHCO₃, dried, concentrated to afford the impure product (590mg, yield: 93%). 150 mg of the impure product was purified via pre-HPLCto obtain the final product (80 mg, yield: 60%).

¹H NMR (400 MHz, CDCl₃): 7.68 (s, 1H); 7.37-7.24 (m, 5H); 3.76 (s, 2H);3.20 (s, 2H); 3.18 (d, J=8.8 Hz, 2H); 2.32 (d, J=6.0 Hz, 2H); 2.14-2.06(m, 4H); 1.71 (d, J=10.0 Hz, 2H); 0.89-0.86 (m, 1H); 0.56-0.51 (m, 2H);0.14-0.10 (m, 2H). LCMS (ESI) calcd for C₁₈H₂₅N₃O (m/z): 299.41. found:300.5 [M+1]+.

Preparation of Compound C0126M

a. Preparation of Compound C0126M-1

A mixture of 4-(cyclopropylmethyl)-cyclohexanone (1 g, 6.5 mmol) andethanolamine (4 mL, 65 mmol) in CH₃CH₂OH (20 mL) was stirred overnight(about 18 hours) at room temperature. The mixture was concentrated toremove the solvent. The residue was dissolved in CH₂Cl₂ and washed withsaturated aqueous of Na₂CO₃, dried, and concentrated to afford the finalproduct without any purification (1.06 g, Yield: 84%). The product'sidentity was confirmed by ¹H NMR and MS (target M+1=197).

b. Preparation of Compound C0126M

A mixture of C0126M-1 (122 mg, 0.63 mmol) and 4-(methylsulfonyl)benzene-1-sulfonyl chloride (200 mg, 0.69 mmol) intriethylamine (0.1 mL, 0.69 mmol) and CHCl₃ (2.4 mL) was stirredovernight (about 18 hours) at room temperature. The solvent was removed.The residue was washed with diethyl ether (3×) and the 1N HCl was addedto the residue until pH=1. The mixture wad washed with diethyl ether(2×). Then 1N NaOH was added until a precipitate formed (pH=12). Thesolid was filtered and dissolved in CHCl₃, dried with Na₂SO₄, andconcentrated to afford the final product (28 mg, Yield: 11%). Thestructure was confirmed by ¹H NMR & MS (target: M+1=415), purity 95.3%by HPLC.

¹H NMR (400 MHz, CDCl₃): 8.07 (s, 4H); 3.97 (t, J=6.4 Hz, 2H); 3.55 (t,J=6.4 Hz, 2H); 3.09 (s, 3H); 2.99 (dd, J=9.2 Hz, 2.4 Hz, 2H); 2.51 (td,J=12.8 Hz, 4 Hz, 2H); 2.27-2.19 (m, 4H); 1.56 (d, J=11.6 Hz, 2H);0.87-0.84 (m, 1H); 0.52-0.48 (m, 2H); 0.09-0.07 (m, 2H). MS (ESI) calcdfor C₁₈H₂₆N₂O₅S₂ (m/z): 414.54. found: 415.3 [M+1]⁺.

Preparation of Compound C0128M

a. Preparation of Compound C0128M-1

Compound C0065-3 (213 mg, 1.8 mmol) was addedN-(cyclopropylmethyl)-piperidine-4-one (306 mg, 2.1 mmol) in ethanol (2mL). The mixture was stirred overnight (about 18 hours) at roomtemperature. The solvent was removed under reduced pressure to give thecrude product as yellow oil. The crude product was used for next stepwithout any further purification (519 mg; yield: 100%).

b. Preparation of Compound C0128M

To a solution of compound C0128M-1 (456 mg, 1.8 mmol) in pyridine (2 mL)was added 4-acetyl-benzene-1-sulfonyl chloride (473 mg, 2.2 mmol). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. The residue was diluted withCH₂Cl₂ (100 mL) and washed with saturated sodium carbonate solution (50mL×3). Then the organic layer was dried and concentrated to get thecrude product as dark oil (700 mg).

¹H NMR (400 MHz, CDCl₃): 8.05 (d, J=8.8 Hz, 2H); 7.97 (d, J=8.8 Hz, 2H);4.26-4.19 (m, 1H); 3.76 (dd, J=8.7, 5.6 Hz, 1H); 3.11 (t, J=8.8 Hz, 1H);2.97 (m, 2H); 2.66 (s, 3H); 2.61 (dd, J=12.8, 4.4 Hz, 1H); 2.45 (dd,J=5.6, 1.5 Hz, 2H); 2.39 (dd, J=13.0, 4.4 Hz, 1H); 2.26-2.15 (m, 10H);1.75 (m, 1H); 1.44-1.37 (m, 1H); 0.86 (m, 1H); 0.54-0.45 (m, 2H); 0.09(m, 2H); LCMS (ESI) calcd for C₂₂H₃₃N₃O₄S (m/z): 435.58. found: 436.4[M+1]⁺.

Preparation of Compound C0129M

a. Preparation of Compound C0129M-1

Run 1: To a solution of compound C0084M-2 (617 mg, 4.0 mmol) in methanol(15 mL) was added 1-(cyclopropylmethyl)piperidin-4-one (307 mg, 2.0mmol). The mixture was stirred at 70° C. overnight. The solvent wasremoved under reduced pressure. The residue was purified by columnchromatography (dichloromethane:methanol=3:1) to obtain the unknowncompound as yellow solid (300 mg).

Run 2: To a solution of compound C0084M-2 (309 mg, 2.0 mmol) in methanol(15 mL) was added 1-(cyclopropylmethyl)piperidin-4-one (341 mg, 2.2mmol) and 4-methylbenzenesulfonic acid monohydrate (12 mg, 0.06 mmol).The mixture was stirred at room temperature overnight (about 18 hours).Then the reaction was refluxed for 7 hours. The solvent was removedunder reduced pressure. The residue was purified by columnchromatography (dichloromethane:methanol=3:1) to obtain the unknowncompound as yellow solid (120 mg).

Preparation of Compound C0133M

Prepared using methods discussed elsewhere herein.

¹H NMR (400 MHz, CDCl₃): 7.80 (d, J=9.2 Hz, 2H); 6.95 (d, J=8.8 Hz, 2H);3.93 (t, J=5.6 Hz, 2H); 3.86 (s, 3H); 3.48 (t, J=6.8 Hz, 2H); 2.74 (m,3H); 2.47-2.43 (m, 2H); 2.37-2.31 (m, 2H); 1.59 (d, J=11.6 Hz, 2H); 1.01(d, J=6.0 Hz, 6H); MS (ESI) calcd for C₁₇H₂₆N₂O₄S (m/z): 354.16. found:355.2 [M+1]+.

Preparation of Compound C00134M

a. Preparation of Compound C0134M

A mixture of C0134M-1 (2.1 g, 10.7 mmol) and p-toluenesulfonyl chloride(2.2 g, 11.8 mol) in triethylamine (1.64 mL, 11.8 mmol) and CHCl₃ (40mL) was stirred overnight (about 18 hours) at room temperature. Themixture was washed with NaHCO₃, and the organic layer was concentratedto afford the crude product. The crude product was purified via columnchromatography (CH₂Cl₂:CH₃COCH₃=100/1 to 1/1) to obtain the impureproduct as 2 samples. (210 mg, HPLC: 93%, yield: 6%; 400 mg, HPLC: 83%yield: 11%). The impure product was further purified via preparativeHPLC to obtain the final product (130 mg, yield: 3.7%).

¹H NMR (400 MHz, CDCl₃): 7.77 (d, J=8.0 Hz, 2H); 7.30 (d, J=8.0 Hz, 2H);3.94 (t, J=6.4 Hz, 2H); 3.52 (t, J=6.4 Hz, 2H); 2.98 (m, 2H); 2.54 (td,J=13.2, 4.8 Hz, 2H); 2.42 (s, 3H); 2.27-2.21 (m, 4H); 1.58 (d, J=11.2Hz, 2H); 0.89-0.84 (m, 1H); 0.53-0.49 (m, 2H); 0.10-0.04 (m, 2H). MS(ESI) calcd for C₁₈H₂₆N₂O₃S (m/z): 350.48. found: 351.2 [M+1]+.

Preparation of F-C0134 HCl Salt

a. Preparation of Compound C0027-1

2-Aminoethanol (20 g, 327 mmol) was added to a solution ofN-Boc-piperidin-4-one (21.2 g, 109 mmol) in ethanol (200 ml). Themixture was stirred at room temperature overnight (about 18 hours). Thesolvent was removed under reduced pressure. The residue was diluted withCH₂Cl₂ (800 ml) and washed with saturated aqueous Na₂CO₃ (150 mL×6). Theorganic phase was dried over anhydrous Na₂SO₄ and concentrated toprovide the title compound as colorless oil (25 g, yield: 95%).

¹H-NMR (400 MHz, CDCl₃): 3.75 (t, J=6.4 Hz, 2H); 3.56 (m, 2H); 3.43-3.37(m, 2H); 3.17 (t, J=6.4 Hz, 2H); 1.76 (s, 2H); 1.62 (s, 2H); 1.44 (s,9H).

b. Preparation of Compound A-1

4-Fluorobenzene-1-sulfonyl chloride (24.3 g, 125 mmol) was added to asolution of compound C0027-1 (25 g, 104 mmol) in 200 ml of pyridine. Themixture was stirred overnight (about 18 hours) at room temperature. Thereaction mixture was concentrated to remove the pyridine. The residuewas dissolved with dichloromethane and washed with saturated NaHCO₃. Theorganic layer was dried over Na₂SO₄ and concentrated to get the crudeproduct. The crude product was washed with hexane to provide 38 g of thetitle compound as a brown solid (yield: 91%).

¹H-NMR (400 MHz, CDCl₃): 7.88˜7.85 (m, 2H); 7.19 (t, J=8.4 Hz, 2H); 4.09(m, 2H); 3.97 (t, J=6.4 Hz, 2H); 3.50 (m, 2H); 2.88 (m, 2H); 2.35 (m,2H); 1.57 (m, 2H); 1.46 (s, 9H). MS (ESI) calcd for C₁₈H₂₅FN₂O₅S (m/z):400.15. found: 400.9 [M+1]+.

c. Preparation of Compound A-2

CF₃COOH (109 mL) was added to a solution of compound A-1 (38 g, 95 mmol)in dichloromethane (250 mL). The mixture was stirred for 1.5 hours atroom temperature. 550 ml of dichloromethane was added to the reactionmixture. The mixture was washed with saturated Na₂CO₃, the organic layerdried over Na₂SO₄ and concentrated to get a crude product. The crudeproduct was purified by chromatography to provide compound A-2 as paleyellow oil (13 g, yield: 50%).

¹H-NMR (400 MHz, CDCl₃): 7.91-7.87 (m, 2H); 7.19 (t, J=8.4 Hz, 2H); 3.98(t, J=6.4 Hz, 2H); 3.49 (t, J=6 Hz, 2H); 3.03 (dd, J=12 Hz, 4.8 Hz, 2H);2.83 (td, J=12.4 Hz, 2.0 Hz, 2H); 2.34 (td, J=12.4 Hz, 4.8 Hz, 2H); 1.64(d, J=12 Hz, 2H).

d. Preparation of F-C0134

To a mixture of compound A-2 (13 g, 43 mmol) and K₂CO₃ (23.9 g, 173mmol) in 250 ml of acetone, cyclopropylmethyl bromide (5.85 g, 43 mmol)was added. The reaction mixture was stirred at reflux overnight (about18 hours). After cooling, the mixture was filtered and the filtrate wasconcentrated to obtain the crude product. The crude product was purifiedby chromatography with ethyl acetate yielding the title product as paleyellow oil (13 g, yield: 84%).

¹H-NMR (400 MHz, CDCl₃): 7.91˜7.88 (m, 2H); 7.19 (t, J=8.4 Hz, 2H); 3.95(t, J=6.4 Hz, 2H); 3.50 (t, J=6.4 Hz, 2H); 2.98-2.95 (m, 2H); 2.51 (td,J=12.8 Hz, 4.8 Hz, 2H); 2.26 (d, J=6.8 Hz, 2H); 2.22˜2.16 (m, 2H); 1.55(d, J=12.4 Hz, 2H); 0.88˜0.82 (m, 1H); 0.50˜0.48 (m, 2H); 0.07 (m, 2H).

e. Preparation of F-C0134 HCl Salt

A solution of compound F-C0134 (13 g, 36.7 mmol) in diethyl ether (260mL) was stirred and cooled at −20° C. by dry ice/acetone bath. Asolution of HCl/(CH₃CH₂)₂O (120 mL) was added. The mixture was stirredat −20° C. for 10 minutes. Then, the precipitate was filtered and washedwith hexane/acetone (5:1) to provide the product as an off-white solid(6.6 g, 47% yield).

¹H-NMR (400 MHz, DMSO-d₆): 10.13 (brs, 1H); 7.96 (m, 2H); 7.49 (t, J=8.4Hz, 2H); 3.99 (t, J=6.4 Hz, 2H); 3.55-3.50 (m, 4H); 3.00-2.93 (m, 4H);2.59 (td, J=13.6 Hz, 4.0 Hz, 2H); 1.80 (d, J=14 Hz, 2H); 1.05 (m, 1H);0.62 (d, J=7.2 Hz, 2H); 0.36 (d, J=5.6 Hz, 2H). MS (ESI) calcd. forC₁₇H₂₄ClFN₂O₃S (m/z): 390.90. found: 355.4 [M-HCl]⁺.

Preparation of Compound C0135M

A mixture of compound C0126M-1 (378 mg, 1.92 mmol) and4-ethylbenzene-1-sulfonyl chloride (433 mg, 2.12 mmol) in triethylamine(0.3 mL, 2.12 mmol) and CHCl₃ (7.6 mL) was stirred overnight (about 18hours) at room temperature. The solvent was removed. The residue wasdried to afford the impure product (630 mg). 360 mg of the impureproduct was purified via preparative thin-layer chromatography (ethylacetate:ethanol=10/1) to afford the final product (25 mg, yield: 6%).

¹H NMR (400 MHz, CDCl₃): 7.79 (d, J=8.8 Hz, 2H); 7.31 (d, J=8.4 Hz, 2H);3.94 (t, J=6.4 Hz, 2H); 3.52 (t, J=6.4 Hz, 2H); 3.00 (d, J=10.4 Hz, 2H);2.70 (q, J=7.6 Hz, 2H); 2.56 (td, J=13.6, 4.4 Hz, 2H); 2.28-2.21 (m,4H); 1.59 (d, J=11.6 Hz, 2H); 1.26 (t, J=7.6 Hz, 3H); 0.91-0.83 (m, 1H);0.52-0.50 (m, 2H); 0.10-0.05 (m, 2H). MS (ESI) calcd for C₁₉H₂₈N₂O₃S(m/z): 364.5. found: 365.2 [M+1]⁺.

Preparation of Compound C0136

a. Preparation of Compound C0136-1

Triethylamine (0.73 mL, 5.25 mmol) was added to the suspension of2-aminoacetamide hydrochloride (553 mg, 5.0 mmol) in 10 mL of methanolat room temperature. Then, 2-tetralone (730 mg, 5.0 mmol) was addeddropwise and the resulting mixture stirred overnight (about 18 hours) atroom temperature. NaBH₄ (345 mg, 7.5 mmol) was added and stirred for 2hours. After 2 hours, a white solid was removed via filtration and thefiltrate was concentrated. Then 10 mL of water were added and thesolution was extracted with ethyl acetate (EA) (20 mL×3). The organicphase was collected and dried over Na₂SO₄. The Na₂SO₄ was removed andethyl acetate was removed via evaporation. 0.5 M HCl was added to theether phase until the aqueous layer was pH=3. The aqueous phase wasextracted with ether. Then 0.5 M NaOH was added to the aqueous phaseuntil pH=8 and the aqueous phase was then extracted with ethyl acetate.The organic layer was collected and dried over Na₂SO₄. The Na₂SO₄ wasremoved and the filtrate was concentrated in vacuo to obtain the titleproduct as purple solid (530 mg, yield: 52%).

b. Preparation of Compound C0136M-2

1.53 g of piperidin-4-one hydrochloride monohydrate was added in 50 mLof CHCl₃ and 4.2 mL of triethylamine was added. The mixture was heatedto reflux for 0.5 hour. Then 1.28 mL of benzenesulfonyl chloride wasadded and the mixture was stirred at 70° C. for 4 hours. The solutionwas diluted with 50 mL of CHCl₃ and washed water (50 mL×3) and brine.The organic layer was dried over Na₂SO₄ and evaporated to dryness toobtain the title product as white solid. (1.76 g, yield: 74%).

c. Preparation of Compound C0136M

Compound C0136M-2 (251 mg, 1.05 mmol) was dissolved in isopropyl alcohol(10 mL) and C0136M-1 (204 mg, 1.00 mmol) was added. The reaction mixturewas stirred at reflux for 3 days. The precipitate that formed wasfiltered and purified by column chromatography with CH₂Cl₂:CH₃OH=50:1 to30:1 to obtain the title product as white solid (60 mg, yield: 13.4%).The structure was confirmed by ¹H NMR and LC-MS, Purity: 95.7% by HPLC,shown as follows:

¹H NMR (400 MHz, CDCl₃): 7.76-7.69 (m, 3H); 7.54-7.51 (m, 3H); 7.15-7.05(m, 3H); 3.88 (d, J=12.0 Hz, 2H); 3.37 (s, 2H); 3.10-3.09 (m, 1H);2.91-2.85 (m, 3H); 2.77-2.72 (m, 1H); 2.42 (t, J=13.2 Hz, 2H); 2.12-2.03(m, 2H); 1.92-1.89 (m, 1H); 1.81-1.78 (m, 1H); 1.68-1.56 (m, 2H); calcdfor C₂₃H₂₇N₃O₃S (m/z): 425.18. found: 426.4 [M+1]⁺.

Preparation of Compound C0137M

a. Preparation of Compound C0137M-1

Piperidin-4-one hydrochloride monohydrate (2.9 g) was added in 50 mL ofacetonitrile and then 7.53 g of K₂CO₃ and 4.28 g of bromocyclohexane wasadded. The mixture was heated to reflux overnight (about 18 hours). Themixture was filtered and the filtrate was evaporated to dryness. Theresidue was diluted with CH₂Cl₂ and washed with water (3×) and brine.The organic layer was dried over Na₂SO₄ and evaporated to dryness toobtain 300 mg of the crude product as yellow solid (yield: 6%). Theresidue was washed with 40 mL of diethyl ether and filtered to obtain150 mg of yellow solid (yield: 3%). The diethyl ether, which was used towash the residue, was evaporated and 150 mg of yellow oil was obtained(yield: 3%)

b. Preparation of Compound C0137M

A solution of C0137M-1 (150 mg, 0.83 mmol,), C0104M-1 (272 mg, 1.66mmol) and p-toluenesulfonic acid monohydrate (8 mg, 0.0415 mmol) inCH₃OH (4 mL) was heated to reflux overnight (about 18 hours). Thereaction solution was cooled to room temperature and the white solid wasfiltered and washed with CH₃OH dried in vacuo to obtain 55 mg of theimpure product as white solid. The impure product was purified bypreparative-HPLC to obtain the title product as white solid (13 mg,yield: 4.8%). The structure was confirmed by ¹H NMR & LC-MS, (targetM+1=328), purity: 98.8%.

¹H NMR (400 MHz, CDCl₃): 7.35-7.29 (m, 5H); 6.92 (s, 1H); 3.75 (s, 2H);3.20 (s, 2H); 3.00 (d, J=11.2 Hz, 2H); 2.30 (m, 3H), 2.04 (dt, J=12.4,4.0 Hz, 2H), 1.86-1.78 (m, 4H), 1.71 (d, J=11.4, 2H), 1.20 (m, 6H); MS(ESI) calcd for C₂₀H₂₉N₃O (m/z): 327.46. found: 328.4 [M+1]⁺.

Preparation of Compound C0141M-2

a. Preparation of Compound C0141M-1

Piperidin-4-one (0.5 g) was dissolved in 25 mL of CHCl₃ and 1.4 mL oftriethylamine was added. Then 915 mg of thiophene-2-sulfonyl chloridewas added and the mixture was stirred at 70° C. overnight (about 18hours). The solution was diluted with 50 mL of CHCl₃ and washed 0.1 MHCl (aq, 50 mL×3) and brine. The organic layer was dried over Na₂SO₄ andevaporated to obtain the title product as yellow solid (1.025 g, yield:83.7%).

b. Preparation of Compound C0141M-2

The solution of C0141M-1 (1.02 g, 4.18 mmol), 2-aminoethanol (4.8 mL,83.6 mmol), and p-toluenesulfonic acid monohydrate (24 mg, 0.125 mmol)in ethanol (30 ml) was stirred at room temperature for 2 days. Thereaction solution was evaporated and the residue was dissolved in CHCl₃and washed with sat.NaHCO₃ (aq) for (6×) and brine. The organic layerwas dried over Na₂SO₄ and evaporated to obtain the title product asyellow oil. (1.175 g, yield: 98%). The structure was confirmed by MS(target M+1=289) and ¹H NMR.

Preparation of Compound C0142

a. Preparation of Compound C0142M-1

Ethyl 3-aminopropanoate hydrochloride (5 g) was added to 50 mL ofsaturated NH₄OH (aq). The mixture was stirred at room temperatureovernight (about 18 hours). Then the mixture was washed withCH₂Cl₂:CH₃OH=10:1 (6×). The resulting water layer was evaporated toobtain the title product as white solid (4.5 g; yield: 98%). Thestructure was confirmed by H NMR and LC-MS (target M+1=89)

b. Preparation of Compound C0142M-2

Piperidin-4-one (250 mg) was dissolved in 15 mL of CHCl₃ and 0.7 mL oftriethylamine was added. Then 522 mg of 4-methoxybenzene-1-sulfonylchloride was added and the mixture was stirred at 70° C. overnight. Thesolution was diluted with 30 mL of CHCl₃ and washed with 0.1 M HCl (aq,30 mL×3) and brine. The organic layer was dried over Na₂SO₄ andevaporated to obtain the title product as yellow solid (580 mg, yield:85.4%)

c. Preparation of Compound C0142M

The solution of C0142M-1 (540 mg, 2 mmol) and C0142M-2 (352 mg, 4 mmol)in CH₃OH (10 ml) was heated to reflux overnight. The reaction solutionwas cooled to RT and evaporated to dryness. The residue was purified bychromatography eluted with CH₂Cl₂:CH₃OH=50:1 to 10:1 to obtain the titleproduct as white solid (32 mg, yield: 5%). The structure was confirmedby NMR & LC-MS (target M+1=340), purity: 98.2% by HPLC.

¹H NMR (400 MHz, CDCl₃): 7.70 (d, J=8.8 Hz, 2H); 7.01 (d, J=8.8 Hz, 2H);6.10 (s, 1H); 3.89 (s, 3H); 3.46 (d, J=11.6 Hz, 2H); 2.99 (t, J=6.0 Hz,2H); 2.80 (t, J=9.8 Hz, 2H); 2.28 (t, J=6.0 Hz, 2H); 1.87-1.78 (m, 4H);MS (ESI) calcd for C₁₅H₂₁N₃O₄S (m/z): 339.13. found: 340.4 [M+1]⁺.

Preparation of Compound C0143M

a. Preparation of Compound C0143M-1

8-Quinolinesulfonyl chloride (1.1 g, 4.8 mmol) and triethylamine (808mg, 8 mmol) was added to a solution of piperidin-4-one (400 mg, 4 mmol)in CHCl₃ (20 mL). The mixture was stirred at room temperature overnight(about 18 hours). The mixture was diluted with CHCl₃, and washed withH₂O (10 mL×3) and 0.1N HCl (10 mL×3), dried over Na₂SO₄ and concentratedto give the title product as yellow solid (1.01 g, yield: 87%).

b. Preparation of Compound C0143M-2

p-Toluenesulfonic acid monohydrate (20 mg) and 2-aminoethanol (2.13 g,35 mmol) was added to a solution of compound C0143M-1 (1.01 g, 3.5 mmol)in ethanol (30 mL). The mixture was stirred at room temperatureovernight (about 18 hours). The mixture was concentrated in vacuo todryness; the residue was diluted with dichloromethane, washed withsaturated Na₂CO₃, dried over Na₂SO₄ and concentrated in vacuo to givethe crude product as yellow solid (783 mg, yield: 67.2%).

Preparation of Compound C0144M-2

a. Preparation of Compound C0144M-1

4-Methoxybenzenesulfonyl chloride (453 mg, 2.2 mmol) and triethylamine(404 mg, 4 mmol) were added to a solution of octahydroquinolin-4(1H)-one(306 mg, 2 mmol) in CHCl₃ (15 mL). The mixture was stirred at roomtemperature overnight (about 18 hours). The mixture was diluted withCHCl₃, and washed with H₂O (10 mL×3) and 0.1N HCl (10 mL×3), dried overNa₂SO₄ and concentrated to give the title product as yellow oil (612 mg,yield: 95%).

b. Preparation of Compound C0144M-2

p-Toluenesulfonic acid monohydrate (10 mg) and 2-aminoethanol (1.15 g,18.9 mmol) were added to a solution of compound C0144M-1 (612 mg, 1.89mmol) in ethanol (20 mL). The mixture was stirred at room temperatureovernight (about 18 hours). The mixture was concentrated in vacuo todryness, the residue was diluted with dichloromethane, washed withsaturated Na₂CO₃, dried over Na₂SO₄ and concentrated in vacuo to givethe crude product as yellow gum (556 mg, yield: 80.4%).

Preparation of Compound C0145M

a. Preparation of Compound C0145M-1

A solution of N-Boc-piperidin-4-one (500 mg, 2.67 mmol) and2-aminoethanol (0.55 mL, 9.34 mmol) in ethanol (4 mL) was stirred atroom temperature overnight (about 18 hours). The solvent was removedunder reduced pressure. The residue was diluted with CH₂Cl₂ (50 mL) andwashed with saturated aqueous Na₂CO₃ (30 mL×6). The organic phase wasdried over anhydrous Na₂SO₄, then concentrated to give target product asyellow oil (614 mg; yield: 95%).

b. Preparation of Compound C0145M-2

4-Methoxybenzene-1-sulfonyl chloride (626 mg, 3.04 mmol) andtriethylamine (0.7 mL) were added to a solution of compound C0145M-1(614 mg, 2.53 mmol) in chloroform (5 mL). The mixture was stirred atroom temperature overnight (about 18 hours). The solvent was removedunder reduced pressure. The residue was purified by columnchromatography (petroleum ether:dichloromethane=1:1) to obtain targetproduct as white solid (500 mg; yield: 48%).

c. Preparation of Compound C0145M-3

CF₃COOH (0.6 mL) was added to the solution of compound C0145M-2 (500 mg,1.21 mmol) in 6 mL dichloromethane and the mixture was kept stirring for0.5 hour at room temperature. Then CH₃CH₂OH/NH₃ (20 mL) was added to thereaction mixture. The solvent was removed under reduced pressure. Theresidue was diluted with dichloromethane (20 mL) and the white solid wasprecipitated. TLC showed that the solid was CF₃COONH₄. Then the mixturewas filtered and the filtrate was concentrated. The residue was purifiedby column chromatography (CH₂Cl₂:CH₃OH=10:1) to obtain the product asyellow solid (290 mg; yield: 76%).

Preparation of Compound C0149M-2

a. Preparation of Compound C0149M-1

4-Phenoxybenzenesulfonyl chloride (500 mg, 1.86 mmol) and triethylamine(404 mg, 4 mmol) were added to a solution of piperidin-4-one (202 mg,2.05 mmol) in CHCl₃ (7 mL). The mixture was stirred at room temperatureovernight (about 18 hours). The mixture was diluted with CHCl₃, andwashed with H₂O (10 mL×3) and 0.1 N HCl (10 mL×3), dried over Na₂SO₄ andconcentrated to give 417 mg of the title product (67.8% yield).

b. Preparation of Compound C0149M-2

To a solution of C0149M-1 (417 mg, 1.26 mmol) in ethanol (15 mL) wasadded p-toluenesulfonic acid monohydrate (10 mg) and 2-aminoethanol (769mg, 12.6 mmol). The mixture was stirred at room temperature overnight(about 18 hours). The mixture was concentrated in vacuo to dryness, theresidue was diluted with CH₂Cl₂, washed with saturated Na₂CO₃, driedover Na₂SO₄ and concentrated in vacuo to give the crude product as whitesolid (434 mg, yield: 92.3%).

Preparation of Compound C0150M-2

a. Preparation of Compound C0150M-1

A solution of piperidin-4-one (47 mg, 0.47 mmol) in chloroform (3 mL)was treated with 1-benzothiophene-3-sulfonyl chloride (100 mg, 0.43mmol) and triethylamine (0.12 mL). The mixture was stirred at roomtemperature overnight (about 18 hours). Then, the solvent was removedunder reduced pressure. The residue was diluted in dichloromethane (30mL). The solution was washed by 0.1 N HCl (30 mL×2) and brine. Theorganic phase was dried over anhydrous Na₂SO₄ and concentrated to obtaintarget product as light yellow solid (120 mg; yield: 94%).

b. Preparation of Compound C0150M-2

A solution of C0150M-1 (217 mg, 0.74 mmol) in ethanol (10 mL) wastreated with p-toluenesulfonic acid monohydrate (6 mg) and2-aminoethanol (452 mg, 7.4 mmol). The mixture was stirred at roomtemperature for 5 hours. The mixture was concentrated in vacuo todryness, the residue was diluted with dichloromethane, washed withsaturated Na₂CO₃, dried over Na₂SO₄ and concentrated in vacuo to givethe crude product as light yellow solid (228 mg, yield: 91.2%)

Preparation of Compound C0151M-2

a. Preparation of Compound C0151M-1

A mixture of piperidin-4-one (643 mg, 6.5 mmol) and1-isocyanato-4-methoxybenzene (500 mg, 3.4 mmol) and K₂CO₃ (1.1 g, 8.0mmol) in CH₂Cl₂ (20 mL) was stirred overnight (about 18 hours) at roomtemperature. To the reaction mixture was added 5 mL water and the waterphase was extracted with CH₂Cl₂. The combined organic layer was washedwith 1 N HCl and brine, dried, and concentrated to obtain the product(800 mg, yield: 96%).

b. Preparation of Compound C0151M-2

To a solution of C0151M-1 (0.5 g, 2.0 mmol) in ethanol (5.0 mL) wasadded ethanolamine (0.4 mL). The mixture was stirred at room temperaturefor 4 hours. Then the solvent was removed under reduced pressure. Theresidue was diluted with CH₂Cl₂ and washed with saturated aqueousNa₂CO₃. The organic phase was dried over anhydrous Na₂SO₄, andconcentrated to obtain the title product as white solid (530 mg, yield:90%).

Preparation of Compound C0152M-4

a. Preparation of Compound C0152M-1

A mixture of aniline (5 g, 53.8 mmol), 3-bromopropanoic acid (8.2 g,53.8 mmol), triethylamine (10.8 g, 107.6 mmol), sodium iodide (0.05 g),and tetrahydrofuran (50 mL) was stirred at reflux overnight (about 18hours). The reaction mixture was cooled down and filtered. The filtratewas concentrated under reduced pressure. The residue was diluted withdichloromethane (DCM) (100 mL) and washed with water once. The organicphase was dried over anhydrous Na₂SO₄ and concentrated. The residue waspurified by column chromatography (CH₂Cl₂:CH₃OH:CH₃COOH=200:1:1) toobtain compound C0152M-1 as brown oil (3 g; yield: 33%)

b. Preparation of Compound C0152M-2

Compound C0152M-1 (1.5 g, 9 mmol) and polyphosphosphoric acid (1.5 g)were heated at 100° C. for 1 hour. After cooling to 0° C., ice-water wasadded into the reaction mixture. The aqueous phase was neutralized to pH7 with saturated aqueous KHCO₃ at 0° C. and basified to pH 12 withsaturated aqueous K₂CO₃. The aqueous layer was extracted withdichloromethane (150 mL×4). The organic layer were dried over anhydrousNa₂SO₄, filtered, and evaporated under reduced pressure to give a crudeproduct which was purified by column chromatography (petroleumether:ethyl acetate=5:1) to give product as yellow oil (94 mg; yield:7.1%)

c. Preparation of Compound C0152M-3

To a solution of compound C0152M-2 (160 mg, 1.08 mmol) in pyridine (5mL) was added 4-methoxy-benzene-1-sulfonyl chloride (159 mg, 0.77 mmol).The mixture was stirred at room temperature overnight (about 18 hours).The solvent was removed under reduced pressure. The residue was dilutedin dichloromethane (30 mL). The solution was washed by 0.1 N HCl (30mL×3) and brine. The organic phase was dried over anhydrous Na₂SO₄ andconcentrated to obtain the crude product. The crude product was purifiedby column chromatography (petroleum ether:ethyl acetate=5:1) to obtainthe title product as white solid (130 mg; yield: 64%)

d. Preparation of Compound C0152M-4

To a solution of C0152M-3 (170 mg, 0.536 mmol) and 2-aminoethanol (327mg, 5.36 mmol) in ethanol (10 mL) was add 4-methylbenzenesulfonic acidmonohydrate (3 mg, 0.016 mmol). The reaction was stirred at roomtemperature overnight (about 18 hours). The solvent was removed underreduced pressure. The residue was diluted with CH₂Cl₂ (50 mL) and washedwith saturated aqueous Na₂CO₃ (30 ml×6). The organic phase was driedover anhydrous Na₂SO₄, then concentrated to give compound C0152M-4 asyellow solid (175 mg; yield: 90%)

Preparation of Compound C0153M

a. Preparation of Compound C0153M-1

t-Butyl-3-oxopiperidine-1-carboxylate (560 mg, 2.8 mmol) and2-aminoethanol (599 mg, 9.8 mmol) in ethanol (4 mL) were stirred at roomtemperature overnight (about 18 hours). The reaction solution wasevaporated and the residue was dissolved in dichloromethane and washedwith saturated Na₂CO₃ (aq) (6×) and brine. The organic layer was driedover Na₂SO₄ and evaporated to obtain the title product as yellow oil(640 mg, yield: 94%).

b. Preparation of Compound C0153M-2

233 mg of C0153M-1 was dissolved in 1 mL of pyridine. A solution of 222mg of 4-hydroxybenzene-1-sulfonyl chloride in 0.5 mL of pyridine wasadded and the mixture was stirred at room temperature overnight (about18 hours).

Preparation of Compound C0153M-6

a. Preparation of Compound C0153M-4

A solution of tert-butyl 3-oxopiperidine-1-carboxylate (500 mg, 2.51mmol) in HCl/CH₃OH (12.5 mL) was heated to reflux for 0.5 hours. Thesolution was cooled to room temperature and evaporated to dryness toobtain the HCl salt of the title compound as yellow solid (330 mg,yield: 97%).

b. Preparation of Compound C0153M-5

Piperidin-3-one hydrochloride (330 mg) was added to 10 mL ofacetonitrile. Then, 1.38 g of K₂CO₃ and 855 mg of bromobenzene wereadded to the solution. The mixture was heated at reflux overnight (about18 hours). The mixture was filtered and the filtrate was evaporated todryness. The residue was purified by chromatography eluted withdichloromethane:methanol=30:1 to obtain the title product as yellow oil(260 mg, yield: 58.3%).

c. Preparation of Compound C0153M-6

The solution of C0153M-5 (250 mg, 1.32 mmol), 2-aminoethanol (282 mg,4.62 mmol) in CH₃CH₂OH (3 mL) was stirred at room temperature overnight(about 18 hours). The reaction solution was evaporated to dryness andthe residue was dissolved in dichloromethane and washed with saturatedNa₂CO₃ (aq) (6×) and brine. The organic layer was dried over Na₂SO₄ andevaporated to dryness to obtain the title product as a yellow oil (301mg, yield: 98%).

Each of the patents, patent applications and articles cited herein isincorporated by reference. The use of the article “a” or “an” isintended to include one or more.

The foregoing description and the examples are intended as illustrativeand are not to be taken as limiting. Still other variations within thespirit and scope of this invention are possible and will readily presentthemselves to those skilled in the art.

What is claimed:
 1. A compound that corresponds in structure to FormulaA or a pharmaceutically acceptable salt thereof, optionally includingboth individual enantiomeric forms, a racemate, diastereomers andmixtures thereof

wherein Q is CHR⁹ or C(O); Z is CHR¹⁰ or C(O), but only one of Q and Zis C(O); each of m and n is zero or one and the sum of m+n is 1 or 2; G,P and W are selected from the group consisting of NR²⁰, NR², NR⁷, S andO, where R⁷ and R² are the same or different and are H, C(H)_(v)(D)_(h)where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, and R²⁰ is X-circle A-R¹ as definedhereinafter, with the provisos that i) only one of G, P and W is NR²⁰,ii) one of G, P and W must be NR²⁰, iii) P is NR² when other than NR²⁰,iv) one of G and W is other than NR² or NR⁷ in which R² or R⁷ is H or analiphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) the otherof G and W is NR²⁰, NR², or NR⁷ bonded to a Z or Q, respectively, thatis C(O), and v) P is NR² in which R² is other than—S(O)₂C₁-C₃-hydrocarbyl when (a) the sum of m+n is 1 and the Q or Zpresent is CH₂, (b) the G or W that is not NR²⁰ is O, and (c) R²⁰ is—S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen; X is SO₂,C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O); each of d, e, f and k iseither zero or one and the sum of (d+e+f+k)=2, e is zero when d is zero,and k is zero when f is zero; D and F are the same or different and areCH or CD; E and K are the same or different and are CH₂, CHD or CD₂;circle A is an aromatic or heteroaromatic ring system containing onering or two fused rings; R¹ represents up to three substituent groupsthat themselves are the same or different, R^(1a), R^(1b), and R^(1c),wherein each of those three groups, R^(1a-c), is separately selectedfrom the group consisting of H, C₁-C₆ hydrocarbyl, C₁-C₆ hydrocarbyloxy,C₁-C₆ hydrocarbyloxycarbonyl, trifluoromethyl, trifluoromethoxy, C₁-C₇hydrocarboyl, hydroxy-, trifluoromethyl- or halogen-substituted C₁-C₇hydrocarboyl, C₁-C₆ hydrocarbylsulfonyl, C₁-C₆ hydrocarbyloxysulfonyl,halogen, nitro, phenyl, cyano, carboxyl, C₁-C₇ hydrocarbyl carboxylate,carboxamide or sulfonamide wherein the amido nitrogen in either grouphas the formula NR³R⁴ wherein R³ and R⁴ are the same or different andare H, C₁-C₄ hydrocarbyl, or R³ and R⁴ together with the depictednitrogen form a 5-7-membered ring that optionally contains 1 or 2additional hetero atoms that independently are nitrogen, oxygen orsulfur, MAr, where M is —CH₂—, —O— or —N═N— and Ar is a single-ringedaryl or heteroaryl group, and NR⁵R⁶ wherein R⁵ and R⁶ are the same ordifferent and are H, C₁-C₄ hydrocarbyl, C₁-C₄ acyl, C₁-C₄hydrocarbylsulfonyl, or R⁵ and R⁶ together with the depicted nitrogenform a 5-7-membered ring that optionally contains 1 or 2 additionalhetero atoms that independently are nitrogen, oxygen or sulfur; R⁸, R⁹,and R¹⁰ are each H, or two of R⁸, R⁹, and R¹⁰ are H and one is a C₁-C₈hydrocarbyl group that is unsubstituted or is substituted with up tothree atoms that are the same or different and are oxygen or nitrogenatoms; R¹¹, R¹², R¹³ and R¹⁴ are all H, or R¹¹ and R¹³ are H and R¹² andR¹⁴ are H or D, or one of the pair R¹¹ and R¹² or the pair R¹³ and R¹⁴together with the depicted ring form a saturated or unsaturated6-membered ring, and the other pair are each H or they are H and D asrecited herein.
 2. The compound or its pharmaceutically acceptable saltaccording to claim 1, wherein said compound corresponds in structure toa formula selected from the group consisting of:


3. The compound or its pharmaceutically acceptable salt according toclaim 1 that corresponds in structure to Formula B, optionally includingboth individual enantiomeric forms, a racemate, diastereomers andmixtures thereof

wherein G and W are selected from the group consisting of NR²⁰, NR⁷, Sand O, where R² and R⁷ are the same or different and are H,C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, and R²⁰ is X-circle A-R¹, with theprovisos that: i) only one of G or W is NR²⁰, ii) that one of G and Wmust be NR²⁰, iii) the G or W that is not NR²⁰ is other than NR² or NR⁷in which R² or R⁷ is H or an aliphatic C₁ hydrocarbyl when (a) the sumof m+n is 1 and (b) the G or W that is NR²⁰ is bonded to a Z or Q,respectively, that is C(O), and iv) R² of the depicted NR² is other than—S(O)₂C₁-C₃-hydrocarbyl when (a) the sum of m+n is 1 and the Q or Zpresent is CH₂, (b) the G or W that is not NR²⁰ is O, and (c) R²⁰ is—S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen.
 4. Thecompound or its pharmaceutically acceptable salt according to claim 3,wherein said compound corresponds in structure to Formula I

wherein D and F are the same or different and are CH₂, CHD or CD₂, andW, X, Z, Q, n, m, circle A, R¹, R², R⁸ and the R groups therein definedare as described previously, except that i) R² of the depicted NR² isother than —S(O)₂C₁-C₃-hydrocarbyl when (a) the sum of m+n is 1 and theQ or Z present is CH₂, (b) the G or W that is not NR²⁰ is O, and (c) R²⁰is —S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen, andii) W is other than NR² or NR⁷ in which R² or R⁷ is H or an aliphatic C₁hydrocarbyl when (a) the sum of m+n is 1 and (b) Z is C(O).
 5. Thecompound or its pharmaceutically acceptable salt according to claim 4,wherein X is C(O), CH₂, CD₂, or SO₂.
 6. The compound or itspharmaceutically acceptable salt according to claim 4, wherein W is NR⁷,S or O.
 7. The compound or its pharmaceutically acceptable saltaccording to claim 4, wherein circle A is selected from the groupconsisting of phenyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl,triazinyl (1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl),furanyl, thienyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl,naphthyl, benzofuranyl, isobenzofuranyl, benzothiophenyl,isobenzothiophenyl, benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl,quinazolyl, cinnolinyl, quinoxalinyl, naphthyridinyl, andbenzopyrimidinyl.
 8. The compound or its pharmaceutically acceptablesalt according to claim 4, wherein one of Q and Z is CH₂ and the otheris absent.
 9. The compound or its pharmaceutically acceptable saltaccording to claim 4, wherein said compound corresponds in structure toa formula selected from the group consisting of:


10. The compound or its pharmaceutically acceptable salt according toclaim 3, wherein said compound corresponds in structure to Formula II

wherein D and F are the same or different and are CH₂, CHD or CD₂, andX, W, circle A, R¹, R² and the R groups therein defined are as describedpreviously, except that R² of the depicted NR² is other than—S(O)₂C₁-C₃-hydrocarbyl when W is O, and X-circle A-R¹ is—S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen.
 11. Thecompound or its pharmaceutically acceptable salt according to claim 10,wherein X is C(O), CH₂, CD₂, or SO₂.
 12. The compound or itspharmaceutically acceptable salt according to claim 10, wherein W isNR⁷, S or O.
 13. The compound or its pharmaceutically acceptable saltaccording to claim 10, wherein circle A is selected from the groupconsisting of phenyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl,triazinyl (1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl),furanyl, thienyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl,naphthyl, benzofuranyl, isobenzofuranyl, benzothiophenyl,isobenzothiophenyl, benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl,quinazolyl, cinnolinyl, quinoxalinyl, naphthyridinyl, andbenzopyrimidinyl.
 14. The compound or its pharmaceutically acceptablesalt according to claim 10, wherein one of D and F is CH₂.
 15. Thecompound or its pharmaceutically acceptable salt according to claim 10,wherein said compound corresponds in structure to a formula selectedfrom the group consisting of:


16. The compound or its pharmaceutically acceptable salt according toclaim 3, wherein said compound corresponds in structure to Formula III

wherein D and F are the same or different and are CH₂, CHD or CD₂; eachof m and n is one; and W, X, Z, Q, circle A, R¹, R² and the R groupstherein defined are as described previously, except that i) one of Z andQ is C(O), and ii) W is other than NR² or NR⁷ in which R² and R⁷ is H oran aliphatic C₁ hydrocarbyl when Z is C(O).
 17. The compound or itspharmaceutically acceptable salt according to claim 16, wherein X isC(O), CH₂, CD₂, or SO₂.
 18. The compound or its pharmaceuticallyacceptable salt according to claim 16, wherein W is NR⁷, S or O.
 19. Thecompound or its pharmaceutically acceptable salt according to claim 16,wherein circle A is selected from the group consisting of phenyl,pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl(1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl), furanyl,thienyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, naphthyl,benzofuranyl, isobenzofuranyl, benzothiophenyl, isobenzothiophenyl,benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl, quinazolyl,cinnolinyl, quinoxalinyl, naphthyridinyl, and benzopyrimidinyl.
 20. Thecompound or its pharmaceutically acceptable salt according to claim 16,wherein one of Q and Z is C(O) and the other is CH₂.
 21. The compound orits pharmaceutically acceptable salt according to claim 16, wherein saidcompound corresponds in structure to a formula selected from the groupconsisting of:


22. The compound or its pharmaceutically acceptable salt according toclaim 1 that corresponds in structure to Formula C, optionally includingboth individual enantiomeric forms, a racemate, diastereomers andmixtures thereof

wherein G and W are selected from the group consisting of NR², NR⁷, Sand O, where R² and R⁷ are the same or different and are H,C(H)_(v)(D)_(h) where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C_(1l) hydrocarbyl where each of q and r is0, 1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, with the provisos that: i) one of G and Wmust be NR² or NR⁷, and ii) one of G and W is other than NR² or NR⁷ inwhich R² or R⁷ is H or an aliphatic O₁ hydrocarbyl when (a) the sum ofm+n is 1 and (b) the other of G and W is NR² or NR⁷ bonded to a Z or Q,respectively, that is C(O).
 23. The compound or its pharmaceuticallyacceptable salt according to claim 22, wherein said compound correspondsin structure to a formula selected from the group consisting of:


24. The compound or its pharmaceutically acceptable salt according toclaim 22, wherein e and g are both zero.
 25. The compound or itspharmaceutically acceptable salt according to claim 24, wherein saidcompound corresponds in structure to Formula IV

wherein D and F are the same or different and are CH₂, CHD or CD₂; andW, X, Z, g, circle A, R¹, R², R⁸ and the R groups therein defined are asdescribed previously, except that i) W is other than NR² or NR⁷ in whichR² or R⁷ is H or an aliphatic C₁ hydrocarbyl when the sum of m+n is 1and Z is C(O), and ii) R² of the depicted NR² group is other than H oran aliphatic C₁ hydrocarbyl when the sum of m+n is 1, W is NR² or NR⁷,and Q is C(O).
 26. The compound or its pharmaceutically acceptable saltaccording to claim 25, wherein X is C(O), CH₂, CD₂, or SO₂.
 27. Thecompound or its pharmaceutically acceptable salt according to claim 25,wherein W is NR⁷, S or O.
 28. The compound or its pharmaceuticallyacceptable salt according to claim 25, wherein circle A is selected fromthe group consisting of phenyl, pyridyl, pyrazinyl, pyrimidinyl,pyridazinyl, triazinyl (1,3,5-triazinyl, 1,2,4-triazinyl and1,2,3-triazinyl), furanyl, thienyl, oxazolyl, isoxazolyl, thiazolyl,isothiazolyl, naphthyl, benzofuranyl, isobenzofuranyl, benzothiophenyl,isobenzothiophenyl, benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl,quinazolyl, cinnolinyl, quinoxalinyl, naphthyridinyl, andbenzopyrimidinyl.
 29. The compound or its pharmaceutically acceptablesalt according to claim 25, wherein one of Q and Z is C(O) and the otheris CH₂.
 30. The compound or its pharmaceutically acceptable saltaccording to claim 25, wherein said compound corresponds in structure toa formula:


31. The compound or its pharmaceutically acceptable salt according toclaim 22, wherein said compound corresponds in structure to Formula V

wherein D and F are the same or different and are CH₂, CHD or CD₂; andX, W, circle A, R¹, R² and the R groups therein defined are as describedpreviously.
 32. The compound or its pharmaceutically acceptable saltaccording to claim 31, wherein X is C(O), CH₂, CD₂, or SO₂.
 33. Thecompound or its pharmaceutically acceptable salt according to claim 31,wherein W is NR⁷, S or O.
 34. The compound or its pharmaceuticallyacceptable salt according to claim 31, wherein circle A is selected fromthe group consisting of phenyl, pyridyl, pyrazinyl, pyrimidinyl,pyridazinyl, triazinyl (1,3,5-triazinyl, 1,2,4-triazinyl and1,2,3-triazinyl), furanyl, thienyl, oxazolyl, isoxazolyl, thiazolyl,isothiazolyl, naphthyl, benzofuranyl, isobenzofuranyl, benzothiophenyl,isobenzothiophenyl, benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl,quinazolyl, cinnolinyl, quinoxalinyl, naphthyridinyl, andbenzopyrimidinyl.
 35. The compound or its pharmaceutically acceptablesalt according to claim 31, wherein one of D and F is CH₂.
 36. Thecompound or its pharmaceutically acceptable salt according to claim 31,wherein said compound corresponds in structure to a formula:


37. The compound or its pharmaceutically acceptable salt according toclaim 22, wherein said compound corresponds in structure to Formula VI

wherein D and F are the same or different and are CH₂, CHD or CD₂; andeach of m and n is one; W, X, Z, Q, circle A, R¹, R² and the R groupstherein defined are as described previously, and each of m and n is 1,except that i) one of Z and Q is C(O), ii) W is other than NR² or NR⁷ inwhich R² or R⁷ is H or an aliphatic C₁ hydrocarbyl when Z is C(O), andiii) R² of the depicted NR² group is other than H or an aliphatic C₁hydrocarbyl when W is NR² or NR⁷, and Q is C(O).
 38. The compound or itspharmaceutically acceptable salt according to claim 37, wherein X isC(O), CH₂, CD₂, or SO₂.
 39. The compound or its pharmaceuticallyacceptable salt according to claim 37, wherein W is NR⁷, S or O.
 40. Thecompound or its pharmaceutically acceptable salt according to claim 37,wherein circle A is selected from the group consisting of phenyl,pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, triazinyl(1,3,5-triazinyl, 1,2,4-triazinyl and 1,2,3-triazinyl), furanyl,thienyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, naphthyl,benzofuranyl, isobenzofuranyl, benzothiophenyl, isobenzothiophenyl,benzoxazolyl, benzisoxazole, quinolyl, isoquinolyl, quinazolyl,cinnolinyl, quinoxalinyl, naphthyridinyl, and benzopyrimidinyl.
 41. Thecompound or its pharmaceutically acceptable salt according to claim 36,wherein one of Q and Z is C(O) and the other is CH₂.
 42. The compound orits pharmaceutically acceptable salt according to claim 37, wherein saidcompound corresponds in structure to a formula:


43. A pharmaceutical composition comprising an analgesic effectiveamount of a compound of claim 1 dissolved or dispersed in aphysiologically tolerable carrier.
 44. A pharmaceutical compositioncomprising an analgesic effective amount of a compound of claim 2dissolved or dispersed in a physiologically tolerable carrier.
 45. Apharmaceutical composition comprising an analgesic effective amount of acompound of claim 22 dissolved or dispersed in a physiologicallytolerable carrier
 46. A method of reducing one or both of inflammationand pain in a host mammal in need thereof that comprises administeringto that host mammal a pharmaceutical composition containing an analgesiceffective amount of a compound of Formula A or a pharmaceuticallyacceptable salt thereof, optionally including both individualenantiomeric forms, diastereomers and mixtures thereof dissolved ordispersed in a physiologically tolerable carrier

wherein Q is CHR⁹ or C(O); Z is CHR¹⁰ or C(O), but only one of Q and Zis C(O); each of m and n is zero or one and the sum of m+n is 1 or 2; G,P and W are selected from the group consisting of NR²⁰, NR², NR⁷, S andO, where R⁷ and R² are the same or different and are H, C(H)_(v)(D)_(h)where each of v and h is 0, 1, 2 or 3 and v+h=3,C(H)_(q)(D)_(r)-aliphatic C₁-C₁₁ hydrocarbyl where each of q and r is 0,1, or 2 and q+r=0, 1 or 2, aliphatic C₁-C₁₂ hydrocarbyl sulfonyl oraliphatic C₁-C₁₂ hydrocarboyl, and R²⁰ is X-circle A-R¹ as definedhereinafter, with the provisos that i) only one of G, P and W is NR²⁰,ii) one of G, P and W must be NR²⁰, iii) P is NR² when other than NR²⁰,iv) one of G and W is other than NR² or NR⁷ in which R² and R⁷ is H oran aliphatic C₁ hydrocarbyl when (a) the sum of m+n is 1 and (b) theother of G and W is NR²⁰, NR², or NR⁷ bonded to a Z or Q, respectively,that is C(O); and v) P is NR² in which R² is other than—S(O)₂C₁-C₃-hydrocarbyl when (a) the sum of m+n is 1 and the Q or Z thatis present is CH₂, (b) the G or W that is not NR²⁰ is O, and (c) R²⁰ is—S(O)₂-phenyl-R¹, where R¹ is H, C₁-C₃-hydrocarbyl or halogen; X is SO₂,C(O), CH₂, CD₂, OC(O), NHC(NH) or NHC(O); each of d, e, f and k iseither zero or one and the sum of (d+e+f+k)=2, e is zero when d is zero,and k is zero when f is zero; D and F are the same or different and areCH or CD; E and K are the same or different and are CH₂, CHD or CD₂;circle A is an aromatic or heteroaromatic ring system containing onering or two fused rings; R¹ represents up to three substituent groupsthat themselves are the same or different, R^(1a), R^(1b), and R^(1c),wherein each of those three groups, R^(1a-c), is separately selectedfrom the group consisting of H, C₁-C₆ hydrocarbyl, C₁-C₆ hydrocarbyloxy,C₁-C₆ hydrocarbyloxycarbonyl, trifluoromethyl, trifluoromethoxy, C₁-C₇hydrocarboyl, hydroxy-, trifluoromethyl- or halogen-substituted C₁-C₇hydrocarboyl, C₁-C₆ hydrocarbylsulfonyl, C₁-C₆ hydrocarbyloxysulfonyl,halogen, nitro, phenyl, cyano, carboxyl, C₁-C₇ hydrocarbyl carboxylate,carboxamide or sulfonamide wherein the amido nitrogen in either grouphas the formula NR³R⁴ wherein R³ and R⁴ are the same or different andare H, C₁-C₄ hydrocarbyl, or R³ and R⁴ together with the depictednitrogen form a 5-7-membered ring that optionally contains 1 or 2additional hetero atoms that independently are nitrogen, oxygen orsulfur, MAr, where M is —CH₂—, —O— or —N═N— and Ar is a single-ringedaryl or heteroaryl group, and NR⁵R⁶ wherein R⁵ and R⁶ are the same ordifferent and are H, C₁-C₄ hydrocarbyl, C₁-C₄ acyl, C₁-C₄hydrocarbylsulfonyl, or R⁵ and R⁶ together with the depicted nitrogenform a 5-7-membered ring that optionally contains 1 or 2 additionalhetero atoms that independently are nitrogen, oxygen or sulfur; R⁸, R⁹,and R¹⁰ are each H, or two of R⁸, R⁹, and R¹⁰ are H and one is a C₁-C₈hydrocarbyl group that is unsubstituted or is substituted with up tothree atoms that are the same or different and are oxygen or nitrogenatoms; R¹¹, R¹², R¹³ and R¹⁴ are all H, or R¹¹ and R¹³ are H and R¹² andR¹⁴ are H or D, or one of the pair R¹¹ and R¹² or the pair R¹³ and R¹⁴together with the depicted ring form a saturated or unsaturated6-membered ring, and the other pair are each H or they are H and D asrecited herein.
 47. The method according to claim 46, wherein said hostmammal is selected from the group consisting of a primate, a laboratoryrodent, a companion animal, and a food animal.
 48. The method accordingto claim 46, wherein said composition is administered a plurality oftimes over a period of days.
 49. The method according to claim 46,wherein said composition is administered a plurality of times in oneday.
 50. The method according to claim 46, wherein said composition isadministered perorally.
 51. The method according to claim 46, whereinsaid composition is administered parenterally.